bims-nimamd Biomed News
on Neuroimmunity and neuroinflammation in ageing and metabolic disease
Issue of 2022–06–05
107 papers selected by
Fawaz Alzaïd, Sorbonne Université
  1. A cancer-associated RNA polymerase III identity drives robust transcription and expression of snaR-A noncoding RNA.
  2. Artificial neural networks enable genome-scale simulations of intracellular signaling.
  3. ER-lysosome lipid transfer protein VPS13C/PARK23 prevents aberrant mtDNA-dependent STING signaling.
  4. The Ube2m-Rbx1 neddylation-Cullin-RING-Ligase proteins are essential for the maintenance of Regulatory T cell fitness.
  5. Cellular and genetic drivers of RNA editing variation in the human brain.
  6. Mapping the cardiac vascular niche in heart failure.
  7. Disulfiram bolsters T-cell anti-tumor immunity through direct activation of LCK-mediated TCR signaling.
  8. STING agonism reprograms tumor-associated macrophages and overcomes resistance to PARP inhibition in BRCA1-deficient models of breast cancer.
  9. BRD4 promotes resection and homology-directed repair of DNA double-strand breaks.
  10. T1D patient-derived hematopoietic stem cells are programmed to generate Tph, Tfh, and autoimmunity-associated B cell subsets in human immune system mice.
  11. Isolation of Group 2 Innate Lymphoid Cells from Mouse Nasal Mucosa to Detect the Expression of CD226.
  12. Interspecies commensal interactions have nonlinear impacts on host immunity.
  13. Switch Tandem Repeats Influence the Choice of the Alternative End-Joining Pathway in Immunoglobulin Class Switch Recombination.
  14. Induction of synapse formation by de novo neurotransmitter synthesis.
  15. Island-specific evolution of a sex-primed autosome in a sexual planarian.
  16. BCL6-dependent TCF-1+ progenitor cells maintain effector and helper CD4+ T cell responses to persistent antigen.
  17. RNA inhibits dMi-2/CHD4 chromatin binding and nucleosome remodeling.
  18. Characterisation of a nucleo-adhesome.
  19. High-throughput, single-microbe genomics with strain resolution, applied to a human gut microbiome.
  20. Neddylation inhibition induces glutamine uptake and metabolism by targeting CRL3SPOP E3 ligase in cancer cells.
  21. The perception and response of T cells to a changing environment are based on the law of initial value.
  22. Transforming dysfunctional CD8+ T cells into natural controller-like CD8+ T cells: can TCF-1 be the magic wand?
  23. The microRNA cluster C19MC confers differentiation potential into trophoblast lineages upon human pluripotent stem cells.
  24. A multiancestry genome-wide association study of unexplained chronic ALT elevation as a proxy for nonalcoholic fatty liver disease with histological and radiological validation.
  25. Author Correction: Fundamental immune-oncogenicity trade-offs define driver mutation fitness.
  26. Learned iterative segmentation of highly variable anatomy from limited data: Applications to whole heart segmentation for congenital heart disease.
  27. Mitochondrial Exhaustion of Memory CD4 T-Cells in Treated HIV-1 Infection.
  28. Genome-wide protein-DNA interaction site mapping in bacteria using a double-stranded DNA-specific cytosine deaminase.
  29. High heritability of ascending aortic diameter and trans-ancestry prediction of thoracic aortic disease.
  30. The epigenetic regulation of the germinal center response.
  31. Single-Cell Transcriptomic Atlas of Gingival Mucosa in Type 2 Diabetes.
  32. P-selectin mobility undergoes a sol-gel transition as it diffuses from exocytosis sites into the cell membrane.
  33. A calcium-based plasticity model for predicting long-term potentiation and depression in the neocortex.
  34. Sustained Trem2 stabilization accelerates microglia heterogeneity and Aβ pathology in a mouse model of Alzheimer's disease.
  35. Author Correction: The Oligodendrocyte Transcription Factor 2 OLIG2 regulates transcriptional repression during myelinogenesis in rodents.
  36. Reversal of synapse loss in Alzheimer mouse models by targeting mGluR5 to prevent synaptic tagging by C1Q.
  37. Stromal changes in the aged lung induce an emergence from melanoma dormancy.
  38. Membrane curvature regulates the spatial distribution of bulky glycoproteins.
  39. Glucagon-receptor-antagonism-mediated β-cell regeneration as an effective anti-diabetic therapy.
  40. Chemokines form nanoparticles with DNA and can superinduce TLR-driven immune inflammation.
  41. PRODH2-Mediated Proline Metabolism Boosts CAR T-cell Effector Function.
  42. Antibody homotypic interactions are encoded by germline light chain complementarity determining region 2.
  43. Direct interaction of a chaperone-bound type III secretion substrate with the export gate.
  44. α Cell dysfunction in islets from nondiabetic, glutamic acid decarboxylase autoantibody-positive individuals.
  45. Hierarchical micro/nanostructured silver hollow fiber boosts electroreduction of carbon dioxide.
  46. Axonal transport of Hrs is activity dependent and facilitates synaptic vesicle protein degradation.
  47. Dual inhibition of innate immunity and apoptosis by human cytomegalovirus protein UL37x1 enables efficient virus replication.
  48. Cryo-EM structures of human A2ML1 elucidate the protease-inhibitory mechanism of the A2M family.
  49. Kinase-deficient BTK mutants confer ibrutinib resistance through activation of the kinase HCK.
  50. Cell identity conversion in liver regeneration after injury.
  51. Long-term hepatitis B virus infection of rhesus macaques requires suppression of host immunity.
  52. Purine nucleoside phosphorylase enables dual metabolic checkpoints that prevent T cell immunodeficiency and TLR7-dependent autoimmunity.
  53. A fungal dioxygenase CcTet serves as a eukaryotic 6mA demethylase on duplex DNA.
  54. Neuroendocrinal and molecular basis of flight performance in locusts.
  55. LL-37 transports immunoreactive cGAMP to activate STING signaling and enhance interferon-mediated host antiviral immunity.
  56. Stochastic expression of invasion genes in Plasmodium falciparum schizonts.
  57. CD4 T cells are rapidly depleted from tuberculosis granulomas following acute SIV co-infection.
  58. Helix-Loop-Helix Proteins in Adaptive Immune Development.
  59. Islet cell replacement and transplantation immunology in a mouse strain with inducible diabetes.
  60. De novo biosynthesis of rubusoside and rebaudiosides in engineered yeasts.
  61. Spatiotemporal dynamics of noradrenaline during learned behaviour.
  62. Quaternary structure independent folding of voltage-gated ion channel pore domain subunits.
  63. Molecularly defined circuits for cardiovascular and cardiopulmonary control.
  64. Redox regulation by TXNRD3 during epididymal maturation underlies capacitation-associated mitochondrial activity and sperm motility in mice.
  65. Regulation of Hepatic Lipid and Glucose Metabolism by INSP3R1.
  66. Combination anti-HIV antibodies provide sustained virological suppression.
  67. Disrupted control of origin activation compromises genome integrity upon destabilization of Polε and dysfunction of the TRP53-CDKN1A/P21 axis.
  68. Lung Cancer Induces NK Cell Contractility and Cytotoxicity Through Transcription Factor Nuclear Localization.
  69. POWR1 is a domestication gene pleiotropically regulating seed quality and yield in soybean.
  70. Microbiota and adipocyte mitochondrial damage in type 2 diabetes are linked by Mmp12+ macrophages.
  71. Intrinsic antiviral immunity of barrier cells revealed by an iPSC-derived blood-brain barrier cellular model.
  72. Epigenetic Regulation to Enhance Graft-versus-Leukemia Activity.
  73. The genomic landscape of cholangiocarcinoma reveals the disruption of post-transcriptional modifiers.
  74. Common risk variants for epilepsy are enriched in families previously targeted for rare monogenic variant discovery.
  75. Identification of positive and negative regulators of antiviral RNA interference in Arabidopsis thaliana.
  76. A stromal Integrated Stress Response activates perivascular cancer-associated fibroblasts to drive angiogenesis and tumour progression.
  77. The transcription factor Hhex regulates inflammation-related genes in microglia.
  78. Ribosome stalling during selenoprotein translation exposes a ferroptosis vulnerability.
  79. Macrophage Cx43 Is Necessary for Fibroblast Cytosolic Calcium and Lung Fibrosis After Injury.
  80. Human pathogenic RNA viruses establish noncompeting lineages by occupying independent niches.
  81. Leukocyte cytokine responses in adult patients with mitochondrial DNA defects.
  82. Modular (de)construction of complex bacterial phenotypes by CRISPR/nCas9-assisted, multiplex cytidine base-editing.
  83. GPR174 signals via Gαs to control a CD86-containing gene expression program in B cells.
  84. Lifelong changes of neurotransmitter receptor expression and debilitation of hippocampal synaptic plasticity following early postnatal blindness.
  85. The SpoVA membrane complex is required for dipicolinic acid import during sporulation and export during germination.
  86. Mapping cis-regulatory elements in human neurons links psychiatric disease heritability and activity-regulated transcriptional programs.
  87. Fix the process that led to Alzheimer's drug fiasco.
  88. Spatiotemporal multiplexed immunofluorescence imaging of living cells and tissues with bioorthogonal cycling of fluorescent probes.
  89. Shared Clonal Origin of Multiple Histiocytic and Dendritic Neoplasms and Polycythemia Vera, Follicular Lymphoma in 1 Patient.
  90. Sublytic C5b-9 Induces CCL3/4 Production and Macrophage Accumulation in Thy-1N Rats via PKC-α/p65/IRF-8 Axis.
  91. Impact of a 7-day homogeneous diet on interpersonal variation in human gut microbiomes and metabolomes.
  92. Structural variant-based pangenome construction has low sensitivity to variability of haplotype-resolved bovine assemblies.
  93. Long-Term Care for Children With Medical Complexity: A Call for Data.
  94. Cross-tissue, single-cell stromal atlas identifies shared pathological fibroblast phenotypes in four chronic inflammatory diseases.
  95. Myriapod genomes reveal ancestral horizontal gene transfer and hormonal gene loss in millipedes.
  96. Nuclear Vav3 is required for polycomb repression complex-1 activity in B-cell lymphoblastic leukemogenesis.
  97. The nonclassical MHC class I Qa-1 expressed in layer 6 neurons regulates activity-dependent plasticity via microglial CD94/NKG2 in the cortex.
  98. Cryo-EM structures of Gid12-bound GID E3 reveal steric blockade as a mechanism inhibiting substrate ubiquitylation.
  99. A conversation with Elizabeth Blackburn and Viviane Tabar.
  100. Neutralizing Antibodies Against Factor VIII Can Occur Through a Non-Germinal Center Pathway.
  101. Cardiac inducing colonies halt fibroblast activation and induce cardiac/endothelial cells to move and expand via paracrine signaling.
  102. Blocking phospholamban with VHH intrabodies enhances contractility and relaxation in heart failure.
  103. Identification of cell types in multiplexed in situ images by combining protein expression and spatial information using CELESTA.
  104. Role of thymic stromal lymphopoietin in allergy and beyond.
  105. The Shigella Spp. Type III Effector Protein OspB Is a Cysteine Protease.
  106. Genetic correlates of phenotypic heterogeneity in autism.
  107. An integrated model for Gpr124 function in Wnt7a/b signaling among vertebrates.
  1. Nat Commun. 2022 May 30. 13(1): 3007
    A cancer-associated RNA polymerase III identity drives robust transcription and expression of snaR-A noncoding RNA.
    Kevin Van Bortle, David P Marciano, Qing Liu, Tristan Chou, Andrew M Lipchik, Sanjay Gollapudi, Benjamin S Geller, Emma Monte, Rohinton T Kamakaka, Michael P Snyder.
      RNA polymerase III (Pol III) includes two alternate isoforms, defined by mutually exclusive incorporation of subunit POLR3G (RPC7α) or POLR3GL (RPC7β), in mammals. The contributions of POLR3G and POLR3GL to transcription potential has remained poorly defined. Here, we discover that loss of subunit POLR3G is accompanied by a restricted repertoire of genes transcribed by Pol III. Particularly sensitive is snaR-A, a small noncoding RNA implicated in cancer proliferation and metastasis. Analysis of Pol III isoform biases and downstream chromatin features identifies loss of POLR3G and snaR-A during differentiation, and conversely, re-establishment of POLR3G gene expression and SNAR-A gene features in cancer contexts. Our results support a model in which Pol III identity functions as an important transcriptional regulatory mechanism. Upregulation of POLR3G, which is driven by MYC, identifies a subgroup of patients with unfavorable survival outcomes in specific cancers, further implicating the POLR3G-enhanced transcription repertoire as a potential disease factor.
    DOI:  https://doi.org/10.1038/s41467-022-30323-6
  2. Nat Commun. 2022 Jun 02. 13(1): 3069
    Artificial neural networks enable genome-scale simulations of intracellular signaling.
    Avlant Nilsson, Joshua M Peters, Nikolaos Meimetis, Bryan Bryson, Douglas A Lauffenburger.
      Mammalian cells adapt their functional state in response to external signals in form of ligands that bind receptors on the cell-surface. Mechanistically, this involves signal-processing through a complex network of molecular interactions that govern transcription factor activity patterns. Computer simulations of the information flow through this network could help predict cellular responses in health and disease. Here we develop a recurrent neural network framework constrained by prior knowledge of the signaling network with ligand-concentrations as input and transcription factor-activity as output. Applied to synthetic data, it predicts unseen test-data (Pearson correlation r = 0.98) and the effects of gene knockouts (r = 0.8). We stimulate macrophages with 59 different ligands, with and without the addition of lipopolysaccharide, and collect transcriptomics data. The framework predicts this data under cross-validation (r = 0.8) and knockout simulations suggest a role for RIPK1 in modulating the lipopolysaccharide response. This work demonstrates the feasibility of genome-scale simulations of intracellular signaling.
    DOI:  https://doi.org/10.1038/s41467-022-30684-y
  3. J Cell Biol. 2022 Jul 04. pii: e202106046. [Epub ahead of print]221(7):
    ER-lysosome lipid transfer protein VPS13C/PARK23 prevents aberrant mtDNA-dependent STING signaling.
    William Hancock-Cerutti, Zheng Wu, Peng Xu, Narayana Yadavalli, Marianna Leonzino, Arun Kumar Tharkeshwar, Shawn M Ferguson, Gerald S Shadel, Pietro De Camilli.
      Mutations in VPS13C cause early-onset, autosomal recessive Parkinson's disease (PD). We have established that VPS13C encodes a lipid transfer protein localized to contact sites between the ER and late endosomes/lysosomes. In the current study, we demonstrate that depleting VPS13C in HeLa cells causes an accumulation of lysosomes with an altered lipid profile, including an accumulation of di-22:6-BMP, a biomarker of the PD-associated leucine-rich repeat kinase 2 (LRRK2) G2019S mutation. In addition, the DNA-sensing cGAS-STING pathway, which was recently implicated in PD pathogenesis, is activated in these cells. This activation results from a combination of elevated mitochondrial DNA in the cytosol and a defect in the degradation of activated STING, a lysosome-dependent process. These results suggest a link between ER-lysosome lipid transfer and innate immune activation in a model human cell line and place VPS13C in pathways relevant to PD pathogenesis.
    DOI:  https://doi.org/10.1083/jcb.202106046
  4. Nat Commun. 2022 May 31. 13(1): 3021
    The Ube2m-Rbx1 neddylation-Cullin-RING-Ligase proteins are essential for the maintenance of Regulatory T cell fitness.
    Di Wu, Haomin Li, Mingwei Liu, Jun Qin, Yi Sun.
      Neddylation-mediated activation of Cullin-RING E3 Ligases (CRLs) are necessary for the degradation of specific immune regulatory proteins. However, little is known about how these processes govern the function of regulatory T (Treg) cells. Here we show that mice with Treg cell-specific deletion of Rbx1, a dual E3 for both neddylation and ubiquitylation by CRLs, develop an early-onset fatal inflammatory disorder, characterized by disrupted Treg cell homeostasis and suppressive functions. Specifically, Rbx1 is essential for the maintenance of an effector Treg cell subpopulation, and regulates several inflammatory pathways. Similar but less severe phenotypes are observed in mice having Ube2m, a neddylation E2 conjugation enzyme, deleted in their Treg cells. Interestingly, Treg-specific deletion of Rbx2/Sag or Ube2f, components of a similar but distinct neddylation-CRL complex, yields no obvious phenotype. Thus, our work demonstrates that the Ube2m-Rbx1 axis is specifically required for intrinsic regulatory processes in Treg cells; and that Rbx1 might also play Ube2m-independent roles in maintaining the fitness of Treg cells, suggesting a layer of complexity in neddylation-dependent activation of CRLs.
    DOI:  https://doi.org/10.1038/s41467-022-30707-8
  5. Nat Commun. 2022 May 30. 13(1): 2997
    Cellular and genetic drivers of RNA editing variation in the human brain.
    Winston H Cuddleston, Junhao Li, Xuanjia Fan, Alexey Kozenkov, Matthew Lalli, Shahrukh Khalique, Stella Dracheva, Eran A Mukamel, Michael S Breen.
      Posttranscriptional adenosine-to-inosine modifications amplify the functionality of RNA molecules in the brain, yet the cellular and genetic regulation of RNA editing is poorly described. We quantify base-specific RNA editing across three major cell populations from the human prefrontal cortex: glutamatergic neurons, medial ganglionic eminence-derived GABAergic neurons, and oligodendrocytes. We identify more selective editing and hyper-editing in neurons relative to oligodendrocytes. RNA editing patterns are highly cell type-specific, with 189,229 cell type-associated sites. The cellular specificity for thousands of sites is confirmed by single nucleus RNA-sequencing. Importantly, cell type-associated sites are enriched in GTEx RNA-sequencing data, edited ~twentyfold higher than all other sites, and variation in RNA editing is largely explained by neuronal proportions in bulk brain tissue. Finally, we uncover 661,791 cis-editing quantitative trait loci across thirteen brain regions, including hundreds with cell type-associated features. These data reveal an expansive repertoire of highly regulated RNA editing sites across human brain cell types and provide a resolved atlas linking cell types to editing variation and genetic regulatory effects.
    DOI:  https://doi.org/10.1038/s41467-022-30531-0
  6. Nat Commun. 2022 May 31. 13(1): 3027
    Mapping the cardiac vascular niche in heart failure.
    Fabian Peisker, Maurice Halder, James Nagai, Susanne Ziegler, Nadine Kaesler, Konrad Hoeft, Ronghui Li, Eric M J Bindels, Christoph Kuppe, Julia Moellmann, Michael Lehrke, Christian Stoppe, Michael T Schaub, Rebekka K Schneider, Ivan Costa, Rafael Kramann.
      The cardiac vascular and perivascular niche are of major importance in homeostasis and during disease, but we lack a complete understanding of its cellular heterogeneity and alteration in response to injury as a major driver of heart failure. Using combined genetic fate tracing with confocal imaging and single-cell RNA sequencing of this niche in homeostasis and during heart failure, we unravel cell type specific transcriptomic changes in fibroblast, endothelial, pericyte and vascular smooth muscle cell subtypes. We characterize a specific fibroblast subpopulation that exists during homeostasis, acquires Thbs4 expression and expands after injury driving cardiac fibrosis, and identify the transcription factor TEAD1 as a regulator of fibroblast activation. Endothelial cells display a proliferative response after injury, which is not sustained in later remodeling, together with transcriptional changes related to hypoxia, angiogenesis, and migration. Collectively, our data provides an extensive resource of transcriptomic changes in the vascular niche in hypertrophic cardiac remodeling.
    DOI:  https://doi.org/10.1038/s41467-022-30682-0
  7. EMBO J. 2022 May 31. e110636
    Disulfiram bolsters T-cell anti-tumor immunity through direct activation of LCK-mediated TCR signaling.
    Qinlan Wang, Ting Zhu, Naijun Miao, Yingying Qu, Zhuning Wang, Yinong Chao, Jing Wang, Wei Wu, Xinyi Xu, Chenqi Xu, Li Xia, Feng Wang.
      Activation of the T-cell antigen receptor (TCR)-CD3 complex is critical to induce the anti-tumor response of CD8+ T cells. Here, we found that disulfiram (DSF), an FDA-approved drug previously used to treat alcohol dependency, directly activates TCR signaling. Mechanistically, DSF covalently binds to Cys20/Cys23 residues of lymphocyte-specific protein tyrosine kinase (LCK) and enhances its tyrosine 394 phosphorylation, thereby promoting LCK kinase activity and boosting effector T cell function, interleukin-2 production, metabolic reprogramming, and proliferation. Furthermore, our in vivo data revealed that DSF promotes anti-tumor immunity against both melanoma and colon cancer in mice by activating CD8+ T cells, and this effect was enhanced by anti-PD-1 co-treatment. We conclude that DSF directly activates LCK-mediated TCR signaling to induce strong anti-tumor immunity, providing novel molecular insights into the therapeutic effect of DSF on cancer.
    Keywords:  LCK; T cell receptor; cancer immunotherapy; disulfiram; drug repurposing
    DOI:  https://doi.org/10.15252/embj.2022110636
  8. Nat Commun. 2022 May 31. 13(1): 3022
    STING agonism reprograms tumor-associated macrophages and overcomes resistance to PARP inhibition in BRCA1-deficient models of breast cancer.
    Qiwei Wang, Johann S Bergholz, Liya Ding, Ziying Lin, Sheheryar K Kabraji, Melissa E Hughes, Xiadi He, Shaozhen Xie, Tao Jiang, Weihua Wang, Jason J Zoeller, Hye-Jung Kim, Thomas M Roberts, Panagiotis A Konstantinopoulos, Ursula A Matulonis, Deborah A Dillon, Eric P Winer, Nancy U Lin, Jean J Zhao.
      PARP inhibitors (PARPi) have drastically changed the treatment landscape of advanced ovarian tumors with BRCA mutations. However, the impact of this class of inhibitors in patients with advanced BRCA-mutant breast cancer is relatively modest. Using a syngeneic genetically-engineered mouse model of breast tumor driven by Brca1 deficiency, we show that tumor-associated macrophages (TAMs) blunt PARPi efficacy both in vivo and in vitro. Mechanistically, BRCA1-deficient breast tumor cells induce pro-tumor polarization of TAMs, which in turn suppress PARPi-elicited DNA damage in tumor cells, leading to reduced production of dsDNA fragments and synthetic lethality, hence impairing STING-dependent anti-tumor immunity. STING agonists reprogram M2-like pro-tumor macrophages into an M1-like anti-tumor state in a macrophage STING-dependent manner. Systemic administration of a STING agonist breaches multiple layers of tumor cell-mediated suppression of immune cells, and synergizes with PARPi to suppress tumor growth. The therapeutic benefits of this combination require host STING and are mediated by a type I IFN response and CD8+ T cells, but do not rely on tumor cell-intrinsic STING. Our data illustrate the importance of targeting innate immune suppression to facilitate PARPi-mediated engagement of anti-tumor immunity in breast cancer.
    DOI:  https://doi.org/10.1038/s41467-022-30568-1
  9. Nat Commun. 2022 May 31. 13(1): 3016
    BRD4 promotes resection and homology-directed repair of DNA double-strand breaks.
    John K Barrows, Baicheng Lin, Colleen E Quaas, George Fullbright, Elizabeth N Wallace, David T Long.
      Double-strand breaks (DSBs) are one of the most toxic forms of DNA damage and represent a major source of genomic instability. Members of the bromodomain and extra-terminal (BET) protein family are characterized as epigenetic readers that regulate gene expression. However, evidence suggests that BET proteins also play a more direct role in DNA repair. Here, we establish a cell-free system using Xenopus egg extracts to elucidate the gene expression-independent functions of BET proteins in DSB repair. We identify the BET protein BRD4 as a critical regulator of homologous recombination and describe its role in stimulating DNA processing through interactions with the SWI/SNF chromatin remodeling complex and resection machinery. These results establish BRD4 as a multifunctional regulator of chromatin binding that links transcriptional activity and homology-directed repair.
    DOI:  https://doi.org/10.1038/s41467-022-30787-6
  10. Clin Immunol. 2022 May 26. pii: S1521-6616(22)00129-2. [Epub ahead of print] 109048
    T1D patient-derived hematopoietic stem cells are programmed to generate Tph, Tfh, and autoimmunity-associated B cell subsets in human immune system mice.
    Andrea Vecchione, Rachel Madley, Nichole Danzl, Chiara Borsotti, Mohsen Khosravi Marharlooei, Hao-Wei Li, Grace Nauman, Xiaolan Ding, Siu-Hong Ho, Georgia Fousteri, Megan Sykes.
      Interactions between B cells and CD4+ T cells play a central role in the development of Type 1 Diabetes (T1D). Two helper cell subsets, follicular (Tfh) and peripheral (Tph) helper T cells, are increased in patients with T1D but their role in driving B cell autoimmunity is undefined. We used a personalized immune (PI) mouse model to generate human immune systems de novo from hematopoietic stem cells (HSCs) of patients with T1D or from healthy controls (HC). Both groups developed Tfh and Tph-like, and those with T1D-derived immune systems demonstrated increased numbers of Tph-like and Tfh cells compared to HC-derived PI mice. T1D-derived immune systems included increased proportions of unconventional memory CD27-IgD- B cells and reduced proportions of naïve B cells compared to HC PI mice, resembling changes reported for patients with systemic lupus erythematosus. Our findings suggest that T1D HSCs are genetically programmed to produce increased proportions T cells that promote the development of unconventional, possibly autoreactive memory B cells. PI mice provide an avenue for further understanding of the immune abnormalities that drive autoantibody pathogenesis and T1D.
    Keywords:  Autoantibodies; Autoimmunity; Autoreactive; B cell; Hematopoietic stem cells; Human immune system mice; Personalized immune mice; T helper follicular cell; T peripheral helper cell; Type 1 diabetes
    DOI:  https://doi.org/10.1016/j.clim.2022.109048
  11. J Vis Exp. 2022 May 10.
    Isolation of Group 2 Innate Lymphoid Cells from Mouse Nasal Mucosa to Detect the Expression of CD226.
    Yang Xie, Yuan Zhang, Yitian Liu, Yuling Wang, Kun Cheng, Ran Zhuang, Ka Bian.
      With abundant research on group 2 innate lymphoid cells (ILC2s) published over the years, ILC2s are widely known to be implicated in regulating various pathological processes, including anti-helminth immunity, tissue repair, thermogenesis, and autoimmune diseases such as asthma and allergic rhinitis (AR). ILC2s permanently reside in peripheral tissues such as the skin, gut, lungs, and nasal cavity; however, there is limited information about their exact functions in nasal mucosal immunity. CD226 is an activating costimulatory molecule, mainly expressed on natural killer (NK) cells, T cells, and inflammatory monocytes. However, whether ILC2s express CD226 or play a role in the pathogenesis of ILC2s-related diseases remains unknown. Here, we established a method to isolate and identify ILC2s from the nasal mucosa and detected CD226 expression on ILC2s obtained from healthy and AR mice. Herein, we describe this protocol for the isolation and identification of ILC2s from mouse nasal mucosa, which will help explore the internal pathological mechanism of immunological disorders in nasal mucosal diseases.
    DOI:  https://doi.org/10.3791/63525
  12. Cell Host Microbe. 2022 May 24. pii: S1931-3128(22)00264-5. [Epub ahead of print]
    Interspecies commensal interactions have nonlinear impacts on host immunity.
    Tyler A Rice, Agata A Bielecka, Mytien T Nguyen, Connor E Rosen, Deguang Song, Nicole D Sonnert, Yi Yang, Yiyun Cao, Varnica Khetrapal, Jason R Catanzaro, Anjelica L Martin, Saleh A Rashed, Shana R Leopold, Liming Hao, Xuezhu Yu, David van Dijk, Aaron M Ring, Richard A Flavell, Marcel R de Zoete, Noah W Palm.
      The impacts of individual commensal microbes on immunity and disease can differ dramatically depending on the surrounding microbial context; however, the specific bacterial combinations that dictate divergent immunological outcomes remain largely undefined. Here, we characterize an immunostimulatory Allobaculum species from an inflammatory bowel disease patient that exacerbates colitis in gnotobiotic mice. Allobaculum inversely associates with the taxonomically divergent immunostimulatory species Akkermansia muciniphila in human-microbiota-associated mice and human cohorts. Co-colonization with A. muciniphila ameliorates Allobaculum-induced intestinal epithelial cell activation and colitis in mice, whereas Allobaculum blunts the A.muciniphila-specific systemic antibody response and reprograms the immunological milieu in mesenteric lymph nodes by blocking A.muciniphila-induced dendritic cell activation and T cell expansion. These studies thus identify a pairwise reciprocal interaction between human gut bacteria that dictates divergent immunological outcomes. Furthermore, they establish a generalizable framework to define the contextual cues contributing to the "incomplete penetrance" of microbial impacts on human disease.
    Keywords:  IgA; IgA-Seq; gut microbiota; human gut bacteria; human microbiota-associated gnotobiotic mice; immunoglobulin A; immunostimulatory commensals; inflammatory bowel disease; mucosal immunity; reciprocal epistasis
    DOI:  https://doi.org/10.1016/j.chom.2022.05.004
  13. Front Immunol. 2022 ;13 870933
    Switch Tandem Repeats Influence the Choice of the Alternative End-Joining Pathway in Immunoglobulin Class Switch Recombination.
    Chloé Oudinet, Xuefei Zhang, Nadine Puget, Nia Kyritsis, Claire Leduc, Fatima-Zohra Braikia, Audrey Dauba, Frederick W Alt, Ahmed Amine Khamlichi.
      Immunoglobulin class switch recombination (CSR) plays an important role in humoral imm\une responses by changing the effector functions of antibodies. CSR occurs between highly repetitive switch (S) sequences located upstream of immunoglobulin constant gene exons. Switch sequences differ in size, the nature of their repeats, and the density of the motifs targeted by the activation-induced cytidine deaminase (AID), the enzyme that initiates CSR. CSR involves double-strand breaks (DSBs) at the universal Sµ donor region and one of the acceptor S regions. The DSBs ends are fused by the classical non-homologous end-joining (C-NHEJ) and the alternative-NHEJ (A-NHEJ) pathways. Of the two pathways, the A-NHEJ displays a bias towards longer junctional micro-homologies (MHs). The Sµ region displays features that distinguish it from other S regions, but the molecular basis of Sµ specificity is ill-understood. We used a mouse line in which the downstream Sγ3 region was put under the control of the Eµ enhancer, which regulates Sµ, and analyzed its recombination activity by CSR-HTGTS. Here, we show that provision of Eµ enhancer to Sγ3 is sufficient to confer the recombinational features of Sµ to Sγ3, including efficient AID recruitment, enhanced internal deletions and robust donor function in CSR. Moreover, junctions involving Sγ3 display a bias for longer MH irrespective of sequence homology with switch acceptor sites. The data suggest that the propensity for increased MH usage is an intrinsic property of Sγ3 sequence, and that the tandem repeats of the donor site influence the choice of the A-NHEJ.
    Keywords:  B lymphocyte; alternative end-joining; class switch recombination; enhancer; switch sequence
    DOI:  https://doi.org/10.3389/fimmu.2022.870933
  14. Nat Commun. 2022 Jun 01. 13(1): 3060
    Induction of synapse formation by de novo neurotransmitter synthesis.
    Scott R Burlingham, Nicole F Wong, Lindsay Peterkin, Lily Lubow, Carolina Dos Santos Passos, Orion Benner, Michael Ghebrial, Thomas P Cast, Matthew A Xu-Friedman, Thomas C Südhof, Soham Chanda.
      A vital question in neuroscience is how neurons align their postsynaptic structures with presynaptic release sites. Although synaptic adhesion proteins are known to contribute in this process, the role of neurotransmitters remains unclear. Here we inquire whether de novo biosynthesis and vesicular release of a noncanonical transmitter can facilitate the assembly of its corresponding postsynapses. We demonstrate that, in both stem cell-derived human neurons as well as in vivo mouse neurons of purely glutamatergic identity, ectopic expression of GABA-synthesis enzymes and vesicular transporters is sufficient to both produce GABA from ambient glutamate and transmit it from presynaptic terminals. This enables efficient accumulation and consistent activation of postsynaptic GABAA receptors, and generates fully functional GABAergic synapses that operate in parallel but independently of their glutamatergic counterparts. These findings suggest that presynaptic release of a neurotransmitter itself can signal the organization of relevant postsynaptic apparatus, which could be directly modified to reprogram the synapse identity of neurons.
    DOI:  https://doi.org/10.1038/s41467-022-30756-z
  15. Nature. 2022 Jun 01.
    Island-specific evolution of a sex-primed autosome in a sexual planarian.
    Longhua Guo, Joshua S Bloom, Daniel Dols-Serrate, James Boocock, Eyal Ben-David, Olga T Schubert, Kaiya Kozuma, Katarina Ho, Emily Warda, Clarice Chui, Yubao Wei, Daniel Leighton, Tzitziki Lemus Vergara, Marta Riutort, Alejandro Sánchez Alvarado, Leonid Kruglyak.
      The sexual strain of the planarian Schmidtea mediterranea, indigenous to Tunisia and several Mediterranean islands, is a hermaphrodite1,2. Here we isolate individual chromosomes and use sequencing, Hi-C3,4 and linkage mapping to assemble a chromosome-scale genome reference. The linkage map reveals an extremely low rate of recombination on chromosome 1. We confirm suppression of recombination on chromosome 1 by genotyping individual sperm cells and oocytes. We show that previously identified genomic regions that maintain heterozygosity even after prolonged inbreeding make up essentially all of chromosome 1. Genome sequencing of individuals isolated in the wild indicates that this phenomenon has evolved specifically in populations from Sardinia and Corsica. We find that most known master regulators5-13 of the reproductive system are located on chromosome 1. We used RNA interference14,15 to knock down a gene with haplotype-biased expression, which led to the formation of a more pronounced female mating organ. On the basis of these observations, we propose that chromosome 1 is a sex-primed autosome primed for evolution into a sex chromosome.
    DOI:  https://doi.org/10.1038/s41586-022-04757-3
  16. Immunity. 2022 May 25. pii: S1074-7613(22)00223-0. [Epub ahead of print]
    BCL6-dependent TCF-1+ progenitor cells maintain effector and helper CD4+ T cell responses to persistent antigen.
    Yu Xia, Katalin Sandor, Joy A Pai, Bence Daniel, Saravanan Raju, Renee Wu, Sunnie Hsiung, Yanyan Qi, Tenzin Yangdon, Mariko Okamoto, Chun Chou, Kamir J Hiam-Galvez, Robert D Schreiber, Kenneth M Murphy, Ansuman T Satpathy, Takeshi Egawa.
      Soon after activation, CD4+ T cells are segregated into BCL6+ follicular helper (Tfh) and BCL6- effector (Teff) T cells. Here, we explored how these subsets are maintained during chronic antigen stimulation using the mouse chronic LCMV infection model. Using single cell-transcriptomic and epigenomic analyses, we identified a population of PD-1+ TCF-1+ CD4+ T cells with memory-like features. TCR clonal tracing and adoptive transfer experiments demonstrated that these cells have self-renewal capacity and continue to give rise to both Teff and Tfh cells, thus functioning as progenitor cells. Conditional deletion experiments showed Bcl6-dependent development of these progenitors, which were essential for sustaining antigen-specific CD4+ T cell responses to chronic infection. An analogous CD4+ T cell population developed in draining lymph nodes in response to tumors. Our study reveals the heterogeneity and plasticity of CD4+ T cells during persistent antigen exposure and highlights their population dynamics through a stable, bipotent intermediate state.
    Keywords:  CD4(+) T cells; anti-tumor immunity; chronic viral infection; effector T cells; follicular helper T cells; progenitor cells; single-cell genomics
    DOI:  https://doi.org/10.1016/j.immuni.2022.05.003
  17. Cell Rep. 2022 May 31. pii: S2211-1247(22)00670-2. [Epub ahead of print]39(9): 110895
    RNA inhibits dMi-2/CHD4 chromatin binding and nucleosome remodeling.
    Ikram Ullah, Clemens Thölken, Yichen Zhong, Mara John, Oliver Rossbach, Jonathan Lenz, Markus Gößringer, Andrea Nist, Lea Albert, Thorsten Stiewe, Roland Hartmann, Olalla Vázquez, Ho-Ryung Chung, Joel P Mackay, Alexander Brehm.
      The ATP-dependent nucleosome remodeler Mi-2/CHD4 broadly modulates chromatin landscapes to repress transcription and to maintain genome integrity. Here we use individual nucleotide resolution crosslinking and immunoprecipitation (iCLIP) to show that Drosophila Mi-2 associates with thousands of mRNA molecules in vivo. Biochemical data reveal that recombinant dMi-2 preferentially binds to G-rich RNA molecules using two intrinsically disordered regions of unclear function. Pharmacological inhibition of transcription and RNase digestion approaches establish that RNA inhibits the association of dMi-2 with chromatin. We also show that RNA inhibits dMi-2-mediated nucleosome mobilization by competing with the nucleosome substrate. Importantly, this activity is shared by CHD4, the human homolog of dMi-2, strongly suggesting that RNA-mediated regulation of remodeler activity is an evolutionary conserved mechanism. Our data support a model in which RNA serves to protect actively transcribed regions of the genome from dMi-2/CHD4-mediated establishment of repressive chromatin structures.
    Keywords:  ATP-dependent chromatin remodeling; CP: Molecular biology; NuRD; RNA; chromatin; gene regulation; iCLIP
    DOI:  https://doi.org/10.1016/j.celrep.2022.110895
  18. Nat Commun. 2022 Jun 01. 13(1): 3053
    Characterisation of a nucleo-adhesome.
    Adam Byron, Billie G C Griffith, Ana Herrero, Alexander E P Loftus, Emma S Koeleman, Linda Kogerman, John C Dawson, Niamh McGivern, Jayne Culley, Graeme R Grimes, Bryan Serrels, Alex von Kriegsheim, Valerie G Brunton, Margaret C Frame.
      In addition to central functions in cell adhesion signalling, integrin-associated proteins have wider roles at sites distal to adhesion receptors. In experimentally defined adhesomes, we noticed that there is clear enrichment of proteins that localise to the nucleus, and conversely, we now report that nuclear proteomes contain a class of adhesome components that localise to the nucleus. We here define a nucleo-adhesome, providing experimental evidence for a remarkable scale of nuclear localisation of adhesion proteins, establishing a framework for interrogating nuclear adhesion protein functions. Adding to nuclear FAK's known roles in regulating transcription, we now show that nuclear FAK regulates expression of many adhesion-related proteins that localise to the nucleus and that nuclear FAK binds to the adhesome component and nuclear protein Hic-5. FAK and Hic-5 work together in the nucleus, co-regulating a subset of genes transcriptionally. We demonstrate the principle that there are subcomplexes of nuclear adhesion proteins that cooperate to control transcription.
    DOI:  https://doi.org/10.1038/s41467-022-30556-5
  19. Science. 2022 Jun 03. 376(6597): eabm1483
    High-throughput, single-microbe genomics with strain resolution, applied to a human gut microbiome.
    Wenshan Zheng, Shijie Zhao, Yehang Yin, Huidan Zhang, David M Needham, Ethan D Evans, Chengzhen L Dai, Peter J Lu, Eric J Alm, David A Weitz.
      Characterizing complex microbial communities with single-cell resolution has been a long-standing goal of microbiology. We present Microbe-seq, a high-throughput method that yields the genomes of individual microbes from complex microbial communities. We encapsulate individual microbes in droplets with microfluidics and liberate their DNA, which we then amplify, tag with droplet-specific barcodes, and sequence. We explore the human gut microbiome, sequencing more than 20,000 microbial single-amplified genomes (SAGs) from a single human donor and coassembling genomes of almost 100 bacterial species, including several with multiple subspecies strains. We use these genomes to probe microbial interactions, reconstructing the horizontal gene transfer (HGT) network and observing HGT between 92 species pairs; we also identify a significant in vivo host-phage association between crAssphage and one strain of Bacteroides vulgatus. Microbe-seq contributes high-throughput culture-free capabilities to investigate genomic blueprints of complex microbial communities with single-microbe resolution.
    DOI:  https://doi.org/10.1126/science.abm1483
  20. Nat Commun. 2022 May 31. 13(1): 3034
    Neddylation inhibition induces glutamine uptake and metabolism by targeting CRL3SPOP E3 ligase in cancer cells.
    Qiyin Zhou, Wenyu Lin, Chaoqun Wang, Fei Sun, Siwei Ju, Qian Chen, Yi Wang, Yongxia Chen, Haomin Li, Linbo Wang, Zeping Hu, Hongchuan Jin, Xian Wang, Yi Sun.
      Abnormal neddylation activation is frequently observed in human cancers and neddylation inhibition has been proposed as a therapy for cancer. Here, we report that MLN4924, a small-molecule inhibitor of neddylation activating enzyme, increases glutamine uptake in breast cancer cells by causing accumulation of glutamine transporter ASCT2/SLC1A5, via inactivation of CRL3-SPOP E3 ligase. We show the E3 ligase SPOP promotes ASCT2 ubiquitylation, whereas SPOP itself is auto-ubiquitylated upon glutamine deprivation. Thus, SPOP and ASCT2 inversely regulate glutamine uptake and metabolism. SPOP knockdown increases ASCT2 levels to promote growth which is rescued by ASCT2 knockdown. Adding ASCT2 inhibitor V-9302 enhances MLN4924 suppression of tumor growth. In human breast cancer specimens, SPOP and ASCT2 levels are inversely correlated, whereas lower SPOP with higher ASCT2 predicts a worse patient survival. Collectively, our study links neddylation to glutamine metabolism via the SPOP-ASCT2 axis and provides a rational drug combination for enhanced cancer therapy.
    DOI:  https://doi.org/10.1038/s41467-022-30559-2
  21. Sci Signal. 2022 May 31. 15(736): eabj9842
    The perception and response of T cells to a changing environment are based on the law of initial value.
    Eric S Huseby, Emma Teixeiro.
      αβ T cells are critical components of the adaptive immune system and are capable of inducing sterilizing immunity after pathogen infection and eliminating transformed tumor cells. The development and function of T cells are controlled through the T cell antigen receptor, which recognizes peptides displayed on major histocompatibility complex (MHC) molecules. Here, we review how T cells generate the ability to recognize self-peptide-bound MHC molecules and use signals derived from these interactions to instruct cellular development, activation thresholds, and functional specialization in the steady state and during immune responses. We argue that the basic tenants of T cell development and function follow Weber-Fetcher's law of just noticeable differences and Wilder's law of initial value. Together, these laws argue that the ability of a system to respond and the quality of that response are scalable to the basal state of that system. Manifestation of these laws in T cells generates clone-specific activation thresholds that are based on perceivable differences between homeostasis and pathogen encounter (self versus nonself discrimination), as well as poised states for subsequent differentiation into specific effector cell lineages.
    DOI:  https://doi.org/10.1126/scisignal.abj9842
  22. J Clin Invest. 2022 Jun 01. pii: e160474. [Epub ahead of print]132(11):
    Transforming dysfunctional CD8+ T cells into natural controller-like CD8+ T cells: can TCF-1 be the magic wand?
    Hiroshi Takata, Lydie Trautmann.
      HIV infection results in defective CD8+ T cell functions that are incompletely resolved by antiretroviral therapy (ART) except in natural controllers, who have functional CD8+ T cells associated with viral control. In this issue of the JCI, Perdomo-Celis et al. demonstrated that targeting the Wnt/transcription factor T cell factor 1 (Wnt/TCF-1) pathway in dysfunctional CD8+ T cells led to gains in stemness phenotype, metabolic quiescence, survival potential, response to homeostatic γ-chain cytokines, and antiviral capacities, similar to profiles of functional CD8+ T cells in natural controllers. Although reprogramming might not sufficiently reverse the imprinted dysfunction of CD8+ T cells in HIV infection, these findings outline the Wnt/TCF-1 pathway as a potential target to reprogram dysfunctional CD8+ T cells in efforts to achieve HIV remission.
    DOI:  https://doi.org/10.1172/JCI160474
  23. Nat Commun. 2022 Jun 02. 13(1): 3071
    The microRNA cluster C19MC confers differentiation potential into trophoblast lineages upon human pluripotent stem cells.
    Norio Kobayashi, Hiroaki Okae, Hitoshi Hiura, Naoto Kubota, Eri H Kobayashi, Shun Shibata, Akira Oike, Takeshi Hori, Chie Kikutake, Hirotaka Hamada, Hirokazu Kaji, Mikita Suyama, Marie-Line Bortolin-Cavaillé, Jérôme Cavaillé, Takahiro Arima.
      The first cell fate commitment during mammalian development is the specification of the inner cell mass and trophectoderm. This irreversible cell fate commitment should be epigenetically regulated, but the precise mechanism is largely unknown in humans. Here, we show that naïve human embryonic stem (hES) cells can transdifferentiate into trophoblast stem (hTS) cells, but primed hES cells cannot. Our transcriptome and methylome analyses reveal that a primate-specific miRNA cluster on chromosome 19 (C19MC) is active in naïve hES cells but epigenetically silenced in primed ones. Moreover, genome and epigenome editing using CRISPR/Cas systems demonstrate that C19MC is essential for hTS cell maintenance and C19MC-reactivated primed hES cells can give rise to hTS cells. Thus, we reveal that C19MC activation confers differentiation potential into trophoblast lineages on hES cells. Our findings are fundamental to understanding the epigenetic regulation of human early development and pluripotency.
    DOI:  https://doi.org/10.1038/s41467-022-30775-w
  24. Nat Genet. 2022 Jun 02.
    A multiancestry genome-wide association study of unexplained chronic ALT elevation as a proxy for nonalcoholic fatty liver disease with histological and radiological validation.
    Regeneron Genetics Center
      Nonalcoholic fatty liver disease (NAFLD) is a growing cause of chronic liver disease. Using a proxy NAFLD definition of chronic elevation of alanine aminotransferase (cALT) levels without other liver diseases, we performed a multiancestry genome-wide association study (GWAS) in the Million Veteran Program (MVP) including 90,408 cALT cases and 128,187 controls. Seventy-seven loci exceeded genome-wide significance, including 25 without prior NAFLD or alanine aminotransferase associations, with one additional locus identified in European American-only and two in African American-only analyses (P < 5 × 10-8). External replication in histology-defined NAFLD cohorts (7,397 cases and 56,785 controls) or radiologic imaging cohorts (n = 44,289) replicated 17 single-nucleotide polymorphisms (SNPs) (P < 6.5 × 10-4), of which 9 were new (TRIB1, PPARG, MTTP, SERPINA1, FTO, IL1RN, COBLL1, APOH and IFI30). Pleiotropy analysis showed that 61 of 77 multiancestry and all 17 replicated SNPs were jointly associated with metabolic and/or inflammatory traits, revealing a complex model of genetic architecture. Our approach integrating cALT, histology and imaging reveals new insights into genetic liability to NAFLD.
    DOI:  https://doi.org/10.1038/s41588-022-01078-z
  25. Nature. 2022 May 31.
    Author Correction: Fundamental immune-oncogenicity trade-offs define driver mutation fitness.
    David Hoyos, Roberta Zappasodi, Isabell Schulze, Zachary Sethna, Kelvin César de Andrade, Dean F Bajorin, Chaitanya Bandlamudi, Margaret K Callahan, Samuel A Funt, Sine R Hadrup, Jeppe S Holm, Jonathan E Rosenberg, Sohrab P Shah, Ignacio Vázquez-García, Britta Weigelt, Michelle Wu, Dmitriy Zamarin, Laura F Campitelli, Edward J Osborne, Mark Klinger, Harlan S Robins, Payal P Khincha, Sharon A Savage, Vinod P Balachandran, Jedd D Wolchok, Matthew D Hellmann, Taha Merghoub, Arnold J Levine, Marta Łuksza, Benjamin D Greenbaum.
      
    DOI:  https://doi.org/10.1038/s41586-022-04879-8
  26. Med Image Anal. 2022 May 13. pii: S1361-8415(22)00116-5. [Epub ahead of print]80 102469
    Learned iterative segmentation of highly variable anatomy from limited data: Applications to whole heart segmentation for congenital heart disease.
    Danielle F Pace, Adrian V Dalca, Tom Brosch, Tal Geva, Andrew J Powell, Jürgen Weese, Mehdi H Moghari, Polina Golland.
      Training deep learning models that segment an image in one step typically requires a large collection of manually annotated images that captures the anatomical variability in a cohort. This poses challenges when anatomical variability is extreme but training data is limited, as when segmenting cardiac structures in patients with congenital heart disease (CHD). In this paper, we propose an iterative segmentation model and show that it can be accurately learned from a small dataset. Implemented as a recurrent neural network, the model evolves a segmentation over multiple steps, from a single user click until reaching an automatically determined stopping point. We develop a novel loss function that evaluates the entire sequence of output segmentations, and use it to learn model parameters. Segmentations evolve predictably according to growth dynamics encapsulated by training data, which consists of images, partially completed segmentations, and the recommended next step. The user can easily refine the final segmentation by examining those that are earlier or later in the output sequence. Using a dataset of 3D cardiac MR scans from patients with a wide range of CHD types, we show that our iterative model offers better generalization to patients with the most severe heart malformations.
    Keywords:  Congenital heart disease; Interactive segmentation; Recurrent Neural Network; Whole heart segmentation
    DOI:  https://doi.org/10.1016/j.media.2022.102469
  27. Immunometabolism. 2022 ;pii: e220013. [Epub ahead of print]4(2):
    Mitochondrial Exhaustion of Memory CD4 T-Cells in Treated HIV-1 Infection.
    Souheil-Antoine Younes.
      People living with HIV (PLWH) who are immune non-responders (INR) to therapy are unable to restore their CD4 T-cell count and remain at great risk of morbidity and mortality. Here the mitochondrial defects that characterize memory CD4 T-cells in INR and causes of this mitochondrial exhaustion are reviewed. This review also describes the various reagents used to induce the expression of the peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α), the master regulator of mitochondrial biogenesis, which can restore mitochondria fitness and CD4 T-cell proliferation in INR. Due to sustained heightened inflammation in INR, the mitochondrial network is unable to be rejuvenated and requires attenuation of mediators of inflammation to rescue mitochondria and CD4 T-cell counts in INR.
    Keywords:  CD4 T-cells; exhaustion; mitochondria; pgc1α
    DOI:  https://doi.org/10.20900/immunometab20220013
  28. Nat Microbiol. 2022 Jun;7(6): 844-855
    Genome-wide protein-DNA interaction site mapping in bacteria using a double-stranded DNA-specific cytosine deaminase.
    Larry A Gallagher, Elena Velazquez, S Brook Peterson, James C Charity, Matthew C Radey, Michael J Gebhardt, FoSheng Hsu, Lauren M Shull, Kevin J Cutler, Keven Macareno, Marcos H de Moraes, Kelsi M Penewit, Jennifer Kim, Pia A Andrade, Thomas LaFramboise, Stephen J Salipante, Michelle L Reniere, Victor de Lorenzo, Paul A Wiggins, Simon L Dove, Joseph D Mougous.
      DNA-protein interactions are central to fundamental cellular processes, yet widely implemented technologies for measuring these interactions on a genome scale in bacteria are laborious and capture only a snapshot of binding events. We devised a facile method for mapping DNA-protein interaction sites in vivo using the double-stranded DNA-specific cytosine deaminase toxin DddA. In 3D-seq (DddA-sequencing), strains containing DddA fused to a DNA-binding protein of interest accumulate characteristic mutations in DNA sequence adjacent to sites occupied by the DNA-bound fusion protein. High-depth sequencing enables detection of sites of increased mutation frequency in these strains, yielding genome-wide maps of DNA-protein interaction sites. We validated 3D-seq for four transcription regulators in two bacterial species, Pseudomonas aeruginosa and Escherichia coli. We show that 3D-seq offers ease of implementation, the ability to record binding event signatures over time and the capacity for single-cell resolution.
    DOI:  https://doi.org/10.1038/s41564-022-01133-9
  29. Nat Genet. 2022 May 30.
    High heritability of ascending aortic diameter and trans-ancestry prediction of thoracic aortic disease.
    Regeneron Genetics Center
      Enlargement of the aorta is an important risk factor for aortic aneurysm and dissection, a leading cause of morbidity in the developed world. Here we performed automated extraction of ascending aortic diameter from cardiac magnetic resonance images of 36,021 individuals from the UK Biobank, followed by genome-wide association. We identified lead variants across 41 loci, including genes related to cardiovascular development (HAND2, TBX20) and Mendelian forms of thoracic aortic disease (ELN, FBN1). A polygenic score significantly predicted prevalent risk of thoracic aortic aneurysm and the need for surgical intervention for patients with thoracic aneurysm across multiple ancestries within the UK Biobank, FinnGen, the Penn Medicine Biobank and the Million Veterans Program (MVP). Additionally, we highlight the primary causal role of blood pressure in reducing aortic dilation using Mendelian randomization. Overall, our findings provide a roadmap for using genetic determinants of human anatomy to understand cardiovascular development while improving prediction of diseases of the thoracic aorta.
    DOI:  https://doi.org/10.1038/s41588-022-01070-7
  30. Biochim Biophys Acta Gene Regul Mech. 2022 May 25. pii: S1874-9399(22)00043-8. [Epub ahead of print] 194828
    The epigenetic regulation of the germinal center response.
    Shusheng Wu, Yuye Yin, Xiaoming Wang.
      In response to T-cell-dependent antigens, antigen-experienced B cells migrate to the center of the B-cell follicle to seed the germinal center (GC) response after cognate interactions with CD4+ T cells. These GC B cells eventually mature into memory and long-lived antibody-secreting plasma cells, thus generating long-lived humoral immunity. Within GC, B cells undergo somatic hypermutation of their B cell receptors (BCR) and positive selection for the emergence of high-affinity antigen-specific B-cell clones. However, this process may be dangerous, as the accumulation of aberrant mutations could result in malignant transformation of GC B cells or give rise to autoreactive B cell clones that can cause autoimmunity. Because of this, better understanding of GC development provides diagnostic and therapeutic clues to the underlying pathologic process. A productive GC response is orchestrated by multiple mechanisms. An emerging important regulator of GC reaction is epigenetic modulation, which has key transcriptional regulatory properties. In this review, we summarize the current knowledge on the biology of epigenetic mechanisms in the regulation of GC reaction and outline its importance in identification of immunotherapy decision making.
    Keywords:  B cells; Epigenetic modulation; Germinal center response
    DOI:  https://doi.org/10.1016/j.bbagrm.2022.194828
  31. J Dent Res. 2022 Jun 02. 220345221092752
    Single-Cell Transcriptomic Atlas of Gingival Mucosa in Type 2 Diabetes.
    Q Wang, W Lin, X Zhou, K Lei, R Xu, X Zhang, Q Xiong, R Sheng, W Song, W Liu, Q Wang, Q Yuan.
      The oral gingival barrier is a constantly stimulated and dynamic environment where homeostasis is often disrupted, resulting in inflammatory periodontal diseases. Type 2 diabetes (T2D) has been reported to be associated with gingival barrier dysfunction, but the effect and underlying mechanism are inconclusive. Herein, we performed single-cell RNA sequencing (scRNA-seq) of gingiva from leptin receptor-deficient mice (db/db) to examine the gingival heterogeneity in the context of T2D. Periodontal health of control mice is characterized by populations of Krt14+-expressing epithelial cells and Col1a1+-fibroblasts mediating immune homeostasis primarily through the enrichment of innate lymphoid cells. The db/db gingiva exhibited decreased epithelial/stromal ratio and dysfunctional barrier. We further observed stromal, particularly fibroblast immune hyperresponsiveness, linked to the recruitment of myeloid-derived cells at the db/db gingiva. Both scRNA-seq and histological analysis suggested the inflammatory signaling between fibroblasts and neutrophils as a potential driver of diabetes-induced periodontal damage. Notably, the "immune-like" stromal cells were wired toward the induction of gingival γδ T hyperresponsiveness in db/db mice. Our work reveals that the "immune-like" fibroblasts with transcriptional diversity are involved in the innate immune homeostasis at the diabetic gingiva. It highlights a potentially significant role of these cell types in its pathogenesis.
    Keywords:  gingiva barrier; oral mucosal immunity; periodontitis; single-cell RNA sequencing
    DOI:  https://doi.org/10.1177/00220345221092752
  32. Nat Commun. 2022 May 31. 13(1): 3031
    P-selectin mobility undergoes a sol-gel transition as it diffuses from exocytosis sites into the cell membrane.
    Nicola Hellen, Gregory I Mashanov, Ianina L Conte, Sophie le Trionnaire, Victor Babich, Laura Knipe, Alamin Mohammed, Kazim Ogmen, Silvia Martin-Almedina, Katalin Török, Matthew J Hannah, Justin E Molloy, Tom Carter.
      In response to vascular damage, P-selectin molecules are secreted onto the surface of cells that line our blood vessels. They then serve as mechanical anchors to capture leucocytes from the blood stream. Here, we track individual P-selectin molecules released at the surface of live endothelial cells following stimulated secretion. We find P-selectin initially shows fast, unrestricted diffusion but within a few minutes, movement becomes increasingly restricted and ~50% of the molecules become completely immobile; a process similar to a sol-gel transition. We find removal of the extracellular C-type lectin domain (ΔCTLD) and/or intracellular cytoplasmic tail domain (ΔCT) has additive effects on diffusive motion while disruption of the adapter complex, AP2, or removal of cell-surface heparan sulphate restores mobility of full-length P-selectin close to that of ΔCT and ΔCTLD respectively. We have found P-selectin spreads rapidly from sites of exocytosis and evenly decorates the cell surface, but then becomes less mobile and better-suited to its mechanical anchoring function.
    DOI:  https://doi.org/10.1038/s41467-022-30669-x
  33. Nat Commun. 2022 Jun 01. 13(1): 3038
    A calcium-based plasticity model for predicting long-term potentiation and depression in the neocortex.
    Giuseppe Chindemi, Marwan Abdellah, Oren Amsalem, Ruth Benavides-Piccione, Vincent Delattre, Michael Doron, András Ecker, Aurélien T Jaquier, James King, Pramod Kumbhar, Caitlin Monney, Rodrigo Perin, Christian Rössert, Anil M Tuncel, Werner Van Geit, Javier DeFelipe, Michael Graupner, Idan Segev, Henry Markram, Eilif B Muller.
      Pyramidal cells (PCs) form the backbone of the layered structure of the neocortex, and plasticity of their synapses is thought to underlie learning in the brain. However, such long-term synaptic changes have been experimentally characterized between only a few types of PCs, posing a significant barrier for studying neocortical learning mechanisms. Here we introduce a model of synaptic plasticity based on data-constrained postsynaptic calcium dynamics, and show in a neocortical microcircuit model that a single parameter set is sufficient to unify the available experimental findings on long-term potentiation (LTP) and long-term depression (LTD) of PC connections. In particular, we find that the diverse plasticity outcomes across the different PC types can be explained by cell-type-specific synaptic physiology, cell morphology and innervation patterns, without requiring type-specific plasticity. Generalizing the model to in vivo extracellular calcium concentrations, we predict qualitatively different plasticity dynamics from those observed in vitro. This work provides a first comprehensive null model for LTP/LTD between neocortical PC types in vivo, and an open framework for further developing models of cortical synaptic plasticity.
    DOI:  https://doi.org/10.1038/s41467-022-30214-w
  34. Cell Rep. 2022 May 31. pii: S2211-1247(22)00658-1. [Epub ahead of print]39(9): 110883
    Sustained Trem2 stabilization accelerates microglia heterogeneity and Aβ pathology in a mouse model of Alzheimer's disease.
    Rahul Dhandapani, Marilisa Neri, Mario Bernhard, Irena Brzak, Tatjana Schweizer, Stefan Rudin, Stefanie Joller, Ramon Berth, Jasmin Kernen, Anna Neuhaus, Annick Waldt, Rachel Cuttat, Ulrike Naumann, Caroline Gubser Keller, Guglielmo Roma, Dominik Feuerbach, Derya R Shimshek, Ulf Neumann, Fabrizio Gasparini, Ivan Galimberti.
      TREM2 is a transmembrane protein expressed exclusively in microglia in the brain that regulates inflammatory responses to pathological conditions. Proteolytic cleavage of membrane TREM2 affects microglial function and is associated with Alzheimer's disease, but the consequence of reduced TREM2 proteolytic cleavage has not been determined. Here, we generate a transgenic mouse model of reduced Trem2 shedding (Trem2-Ile-Pro-Asp [IPD]) through amino-acid substitution of an ADAM-protease recognition site. We show that Trem2-IPD mice display increased Trem2 cell-surface-receptor load, survival, and function in myeloid cells. Using single-cell transcriptomic profiling of mouse cortex, we show that sustained Trem2 stabilization induces a shift of fate in microglial maturation and accelerates microglial responses to Aβ pathology in a mouse model of Alzheimer's disease. Our data indicate that reduction of Trem2 proteolytic cleavage aggravates neuroinflammation during the course of Alzheimer's disease pathology, suggesting that TREM2 shedding is a critical regulator of microglial activity in pathological states.
    Keywords:  Alzheimer’s disease; Aβ pathology; CP: Neuroscience; Trem2 shedding; Trem2 stabilization; microglia; neuroinflammation; single-cell RNA sequencing
    DOI:  https://doi.org/10.1016/j.celrep.2022.110883
  35. Nat Commun. 2022 Jun 01. 13(1): 3164
    Author Correction: The Oligodendrocyte Transcription Factor 2 OLIG2 regulates transcriptional repression during myelinogenesis in rodents.
    Kunkun Zhang, Shaoxuan Chen, Qihua Yang, Shuanghui Guo, Qiang Chen, Zhixiong Liu, Li Li, Mengyun Jiang, Hongda Li, Jin Hu, Xu Pan, Wenbo Deng, Naian Xiao, Bo Wang, Zhan-Xiang Wang, Liang Zhang, Wei Mo.
      
    DOI:  https://doi.org/10.1038/s41467-022-30945-w
  36. Sci Transl Med. 2022 Jun;14(647): eabi8593
    Reversal of synapse loss in Alzheimer mouse models by targeting mGluR5 to prevent synaptic tagging by C1Q.
    Joshua Spurrier, LaShae Nicholson, Xiaotian T Fang, Austin J Stoner, Takuya Toyonaga, Daniel Holden, Timothy R Siegert, William Laird, Mary Alice Allnutt, Marius Chiasseu, A Harrison Brody, Hideyuki Takahashi, Sarah Helena Nies, Azucena Pérez Cañamás, Pragalath Sadasivam, Supum Lee, Songye Li, Le Zhang, Yiyun H Huang, Richard E Carson, Zhengxin Cai, Stephen M Strittmatter.
      Microglia-mediated synaptic loss contributes to the development of cognitive impairments in Alzheimer's disease (AD). However, the basis for this immune-mediated attack on synapses remains to be elucidated. Treatment with the metabotropic glutamate receptor 5 (mGluR5) silent allosteric modulator (SAM), BMS-984923, prevents β-amyloid oligomer-induced aberrant synaptic signaling while preserving physiological glutamate response. Here, we show that oral BMS-984923 effectively occupies brain mGluR5 sites visualized by [18F]FPEB positron emission tomography (PET) at doses shown to be safe in rodents and nonhuman primates. In aged mouse models of AD (APPswe/PS1ΔE9 overexpressing transgenic and AppNL-G-F/hMapt double knock-in), SAM treatment fully restored synaptic density as measured by [18F]SynVesT-1 PET for SV2A and by histology, and the therapeutic benefit persisted after drug washout. Phospho-TAU accumulation in double knock-in mice was also reduced by SAM treatment. Single-nuclei transcriptomics demonstrated that SAM treatment in both models normalized expression patterns to a far greater extent in neurons than glia. Last, treatment prevented synaptic localization of the complement component C1Q and synaptic engulfment in AD mice. Thus, selective modulation of mGluR5 reversed neuronal gene expression changes to protect synapses from damage by microglial mediators in rodents.
    DOI:  https://doi.org/10.1126/scitranslmed.abi8593
  37. Nature. 2022 Jun 01.
    Stromal changes in the aged lung induce an emergence from melanoma dormancy.
    Mitchell E Fane, Yash Chhabra, Gretchen M Alicea, Devon A Maranto, Stephen M Douglass, Marie R Webster, Vito W Rebecca, Gloria E Marino, Filipe Almeida, Brett L Ecker, Daniel J Zabransky, Laura Hüser, Thomas Beer, Hsin-Yao Tang, Andrew Kossenkov, Meenhard Herlyn, David W Speicher, Wei Xu, Xiaowei Xu, Elizabeth M Jaffee, Julio A Aguirre-Ghiso, Ashani T Weeraratna.
      Disseminated cancer cells from primary tumours can seed in distal tissues, but may take several years to form overt metastases, a phenomenon that is termed tumour dormancy. Despite its importance in metastasis and residual disease, few studies have been able to successfully characterize dormancy within melanoma. Here we show that the aged lung microenvironment facilitates a permissive niche for efficient outgrowth of dormant disseminated cancer cells-in contrast to the aged skin, in which age-related changes suppress melanoma growth but drive dissemination. These microenvironmental complexities can be explained by the phenotype switching model, which argues that melanoma cells switch between a proliferative cell state and a slower-cycling, invasive state1-3. It was previously shown that dermal fibroblasts promote phenotype switching in melanoma during ageing4-8. We now identify WNT5A as an activator of dormancy in melanoma disseminated cancer cells within the lung, which initially enables the efficient dissemination and seeding of melanoma cells in metastatic niches. Age-induced reprogramming of lung fibroblasts increases their secretion of the soluble WNT antagonist sFRP1, which inhibits WNT5A in melanoma cells and thereby enables efficient metastatic outgrowth. We also identify the tyrosine kinase receptors AXL and MER as promoting a dormancy-to-reactivation axis within melanoma cells. Overall, we find that age-induced changes in distal metastatic microenvironments promote the efficient reactivation of dormant melanoma cells in the lung.
    DOI:  https://doi.org/10.1038/s41586-022-04774-2
  38. Nat Commun. 2022 Jun 02. 13(1): 3093
    Membrane curvature regulates the spatial distribution of bulky glycoproteins.
    Chih-Hao Lu, Kayvon Pedram, Ching-Ting Tsai, Taylor Jones, Xiao Li, Melissa L Nakamoto, Carolyn R Bertozzi, Bianxiao Cui.
      The glycocalyx is a shell of heavily glycosylated proteins and lipids distributed on the cell surface of nearly all cell types. Recently, it has been found that bulky transmembrane glycoproteins such as MUC1 can modulate membrane shape by inducing membrane protrusions. In this work, we examine the reciprocal relationship of how membrane shape affects MUC1's spatial distribution on the cell membrane and its biological significance. By employing nanopatterned surfaces and membrane-sculpting proteins to manipulate membrane curvature, we show that MUC1 avoids positively-curved membranes (membrane invaginations) and accumulates on negatively-curved membranes (membrane protrusions). MUC1's curvature sensitivity is dependent on the length and the extent of glycosylation of its ectodomain, with large and highly glycosylated forms preferentially staying out of positive curvature. Interestingly, MUC1's avoidance of positive membrane curvature enables it to escape from endocytosis and being removed from the cell membrane. These findings also suggest that the truncation of MUC1's ectodomain, often observed in breast and ovarian cancers, may enhance its endocytosis and potentiate its intracellular accumulation and signaling.
    DOI:  https://doi.org/10.1038/s41467-022-30610-2
  39. Cell Rep. 2022 May 31. pii: S2211-1247(22)00647-7. [Epub ahead of print]39(9): 110872
    Glucagon-receptor-antagonism-mediated β-cell regeneration as an effective anti-diabetic therapy.
    Yannan Xi, Benbo Song, Iris Ngan, Mark J Solloway, Mark Humphrey, Yan Wang, Kalyani Mondal, Hao Wu, Wenhui Liu, Darrin A Lindhout, Diana Li, Hugo Matern, Avantika Kekatpure, Raj Haldankar, Daniel D Kaplan, Hong Yang, Ophelia Pedersen, Anna Chen, Mei Zhou, Bethany Winans, Wei Guo, Alan Kutach, Marie Fanget, Michael Fox, Jie Tang, Jiping Zha, Husam Younis, David Shen, Alex DePaoli, Hui Tian, Zhonghao Liu.
      Type 1 diabetes mellitus (T1D) is a chronic disease with potentially severe complications, and β-cell deficiency underlies this disease. Despite active research, no therapy to date has been able to induce β-cell regeneration in humans. Here, we discover the β-cell regenerative effects of glucagon receptor antibody (anti-GcgR). Treatment with anti-GcgR in mouse models of β-cell deficiency leads to reversal of hyperglycemia, increase in plasma insulin levels, and restoration of β-cell mass. We demonstrate that both β-cell proliferation and α- to β-cell transdifferentiation contribute to anti-GcgR-induced β-cell regeneration. Interestingly, anti-GcgR-induced α-cell hyperplasia can be uncoupled from β-cell regeneration after antibody clearance from the body. Importantly, we are able to show that anti-GcgR-induced β-cell regeneration is also observed in non-human primates. Furthermore, anti-GcgR and anti-CD3 combination therapy reverses diabetes and increases β-cell mass in a mouse model of autoimmune diabetes.
    Keywords:  CP: Metabolism; anti-diabetic therapy; anti-glucagon receptor; antibody therapy; autoimmune diabetes; beta-cell regeneration; glucagon receptor; type 1 diabetes; α- to β-cell transdifferentiation; β-cell proliferation
    DOI:  https://doi.org/10.1016/j.celrep.2022.110872
  40. J Exp Med. 2022 Jul 04. pii: e20212142. [Epub ahead of print]219(7):
    Chemokines form nanoparticles with DNA and can superinduce TLR-driven immune inflammation.
    Yong Du, Marie Dominique Ah Kioon, Paoline Laurent, Vidyanath Chaudhary, Michael Pierides, Chao Yang, David Oliver, Lionel B Ivashkiv, Franck J Barrat.
      Chemokines control the migratory patterns and positioning of immune cells to organize immune responses to pathogens. However, many chemokines have been associated with systemic autoimmune diseases that have chronic IFN signatures. We report that a series of chemokines, including CXCL4, CXCL10, CXCL12, and CCL5, can superinduce type I IFN (IFN-I) by TLR9-activated plasmacytoid DCs (pDCs), independently of their respective known chemokine receptors. Mechanistically, we show that chemokines such as CXCL4 mediate transcriptional and epigenetic changes in pDCs, mostly targeted to the IFN-I pathways. We describe that chemokines physically interact with DNA to form nanoparticles that promote clathrin-mediated cellular uptake and delivery of DNA in the early endosomes of pDCs. Using two separate mouse models of skin inflammation, we observed the presence of CXCL4 associated with DNA in vivo. These data reveal a noncanonical role for chemokines to serve as nucleic acid delivery vectors to modulate TLR signaling, with implications for the chronic presence of IFN-I by pDCs in autoimmune diseases.
    DOI:  https://doi.org/10.1084/jem.20212142
  41. Cancer Discov. 2022 Jun 02. 12(6): 1405
    PRODH2-Mediated Proline Metabolism Boosts CAR T-cell Effector Function.
      Overexpression of PRODH2 augments CAR T-cell proliferation, effector function, and memory phenotype.
    DOI:  https://doi.org/10.1158/2159-8290.CD-RW2022-054
  42. Proc Natl Acad Sci U S A. 2022 Jun 07. 119(23): e2201562119
    Antibody homotypic interactions are encoded by germline light chain complementarity determining region 2.
    Brandon Leonard, Kannan Sankar, Matthew G Romei, Margaret J Tse, Nina Do, Yanli Yang, Wadim L Matochko, Jack Bevers, Sundeep Bollineni, Kiran Mukhyala, Kam Hon Hoi, Greg A Lazar.
      SignificanceWeak transient interactions are fundamental to immune responses, enabling avidity-driven triggers for pathogen neutralization and cellular regulation. In contrast to obligate binding interactions that can be directly investigated structurally, the low or transitory abundance of weak interactions make them difficult to identify and characterize. This study leverages receptor agonism systems that are sensitive to oligomerization to investigate transient homotypic interfaces between antibody Fab regions. Our results show that self-association determinants are encoded naturally by the antibody germline through light chain complementarity determining region 2 (CDRL2), and these determinants can be engineered into antibodies to enhance their therapeutic properties. Insights into avidity-driven interactions create opportunities for optimization, and accordingly this work expands the engineering toolbox for antibody-based drugs.
    Keywords:  antibody engineering; homotypic interactions; receptor agonism
    DOI:  https://doi.org/10.1073/pnas.2201562119
  43. Nat Commun. 2022 Jun 02. 13(1): 2858
    Direct interaction of a chaperone-bound type III secretion substrate with the export gate.
    Dominic Gilzer, Madeleine Schreiner, Hartmut H Niemann.
      Several gram-negative bacteria employ type III secretion systems (T3SS) to inject effector proteins into eukaryotic host cells directly from the bacterial cytoplasm. The export gate SctV (YscV in Yersinia) binds substrate:chaperone complexes such as YscX:YscY, which are essential for formation of a functional T3SS. Here, we present structures of the YscX:YscY complex alone and bound to nonameric YscV. YscX binds its chaperone YscY at two distinct sites, resembling the heterotrimeric complex of the T3SS needle subunit with its chaperone and co-chaperone. In the ternary complex the YscX N-terminus, which mediates YscX secretion, occupies a binding site within one YscV that is also used by flagellar chaperones, suggesting the interaction's importance for substrate recognition. The YscX C-terminus inserts between protomers of the YscV ring where the stalk protein binds to couple YscV to the T3SS ATPase. This primary YscV-YscX interaction is essential for the formation of a secretion-competent T3SS.
    DOI:  https://doi.org/10.1038/s41467-022-30487-1
  44. J Clin Invest. 2022 Jun 01. pii: e156243. [Epub ahead of print]132(11):
    α Cell dysfunction in islets from nondiabetic, glutamic acid decarboxylase autoantibody-positive individuals.
    HPAP Consortium
      BACKGROUNDMultiple islet autoantibodies (AAbs) predict the development of type 1 diabetes (T1D) and hyperglycemia within 10 years. By contrast, T1D develops in only approximately 15% of individuals who are positive for single AAbs (generally against glutamic acid decarboxylase [GADA]); hence, the single GADA+ state may represent an early stage of T1D.METHODSHere, we functionally, histologically, and molecularly phenotyped human islets from nondiabetic GADA+ and T1D donors.RESULTSSimilar to the few remaining β cells in the T1D islets, GADA+ donor islets demonstrated a preserved insulin secretory response. By contrast, α cell glucagon secretion was dysregulated in both GADA+ and T1D islets, with impaired glucose suppression of glucagon secretion. Single-cell RNA-Seq of GADA+ α cells revealed distinct abnormalities in glycolysis and oxidative phosphorylation pathways and a marked downregulation of cAMP-dependent protein kinase inhibitor β (PKIB), providing a molecular basis for the loss of glucose suppression and the increased effect of 3-isobutyl-1-methylxanthine (IBMX) observed in GADA+ donor islets.CONCLUSIONWe found that α cell dysfunction was present during the early stages of islet autoimmunity at a time when β cell mass was still normal, raising important questions about the role of early α cell dysfunction in the progression of T1D.FUNDINGThis work was supported by grants from the NIH (3UC4DK112217-01S1, U01DK123594-02, UC4DK112217, UC4DK112232, U01DK123716, and P30 DK019525) and the Vanderbilt Diabetes Research and Training Center (DK20593).
    Keywords:  Autoimmune diseases; Diabetes; Endocrinology; Islet cells
    DOI:  https://doi.org/10.1172/JCI156243
  45. Nat Commun. 2022 Jun 02. 13(1): 3080
    Hierarchical micro/nanostructured silver hollow fiber boosts electroreduction of carbon dioxide.
    Shoujie Li, Wei Chen, Xiao Dong, Chang Zhu, Aohui Chen, Yanfang Song, Guihua Li, Wei Wei, Yuhan Sun.
      Efficient conversion of CO2 to commodity chemicals by sustainable way is of great significance for achieving carbon neutrality. Although considerable progress has been made in CO2 utilization, highly efficient CO2 conversion with high space velocity under mild conditions remains a challenge. Here, we report a hierarchical micro/nanostructured silver hollow fiber electrode that reduces CO2 to CO with a faradaic efficiency of 93% and a current density of 1.26 A · cm-2 at a potential of -0.83 V vs. RHE. Exceeding 50% conversions of as high as 31,000 mL · gcat-1 · h-1 CO2 are achieved at ambient temperature and pressure. Electrochemical results and time-resolved operando Raman spectra demonstrate that enhanced three-phase interface reactions and oriented mass transfers synergistically boost CO production.
    DOI:  https://doi.org/10.1038/s41467-022-30733-6
  46. Life Sci Alliance. 2022 Oct;pii: e202000745. [Epub ahead of print]5(10):
    Axonal transport of Hrs is activity dependent and facilitates synaptic vesicle protein degradation.
    Veronica Birdsall, Konner Kirwan, Mei Zhu, Yuuta Imoto, Scott M Wilson, Shigeki Watanabe, Clarissa L Waites.
      Turnover of synaptic vesicle (SV) proteins is vital for the maintenance of healthy and functional synapses. SV protein turnover is driven by neuronal activity in an endosomal sorting complex required for transport (ESCRT)-dependent manner. Here, we characterize a critical step in this process: axonal transport of ESCRT-0 component Hrs, necessary for sorting proteins into the ESCRT pathway and recruiting downstream ESCRT machinery to catalyze multivesicular body (MVB) formation. We find that neuronal activity stimulates the formation of presynaptic endosomes and MVBs, as well as the motility of Hrs+ vesicles in axons and their delivery to SV pools. Hrs+ vesicles co-transport ESCRT-0 component STAM1 and comprise a subset of Rab5+ vesicles, likely representing pro-degradative early endosomes. Furthermore, we identify kinesin motor protein KIF13A as essential for the activity-dependent transport of Hrs to SV pools and the degradation of SV membrane proteins. Together, these data demonstrate a novel activity- and KIF13A-dependent mechanism for mobilizing axonal transport of ESCRT machinery to facilitate the degradation of SV membrane proteins.
    DOI:  https://doi.org/10.26508/lsa.202000745
  47. Nat Microbiol. 2022 May 30.
    Dual inhibition of innate immunity and apoptosis by human cytomegalovirus protein UL37x1 enables efficient virus replication.
    Yujie Ren, An Wang, Di Wu, Chong Wang, Muhan Huang, Xiaobei Xiong, Liang Jin, Wei Zhou, Yang Qiu, Xi Zhou.
      Immune evasion and inhibition of apoptosis are required for successful virus infection. However, inhibition of apoptosis can increase antiviral immune responses, which can then clear viral infections. Here we show that human cytomegalovirus (HCMV)-encoded UL37 exon-1 protein (UL37x1) not only inhibits apoptosis but also suppresses the cGAS-STING immune pathway. Using co-immunoprecipitation assays, we found that UL37x1 binds to TBK1 to abrogate the TBK1-STING-IRF3 interaction. Although the anti-apoptosis function of UL37x1 increases immune signalling, the immunosuppressive role of UL37x1 counteracts this undesirable side-effect. Furthermore, we used mutational analyses to show that the loss of either immunosuppressive or anti-apoptotic function of UL37x1 significantly reduced HCMV replication in human primary foreskin fibroblasts and humanized mice by over twofold. Finally, loss of both functions resulted in over fourfold reduction of HCMV replication in the same cell type and mouse model, showing that both UL37x1 functions are crucial for HCMV infection. We conclude that this sophisticated mechanism enables HCMV to control innate immunity and apoptosis to ensure efficient infection.
    DOI:  https://doi.org/10.1038/s41564-022-01136-6
  48. Nat Commun. 2022 May 31. 13(1): 3033
    Cryo-EM structures of human A2ML1 elucidate the protease-inhibitory mechanism of the A2M family.
    Nadia Sukusu Nielsen, Alessandra Zarantonello, Seandean Lykke Harwood, Kathrine Tejlgård Jensen, Katarzyna Kjøge, Ida B Thøgersen, Leif Schauser, Jesper Lykkegaard Karlsen, Gregers R Andersen, Jan J Enghild.
      A2ML1 is a monomeric protease inhibitor belonging to the A2M superfamily of protease inhibitors and complement factors. Here, we investigate the protease-inhibitory mechanism of human A2ML1 and determine the structures of its native and protease-cleaved conformations. The functional inhibitory unit of A2ML1 is a monomer that depends on covalent binding of the protease (mediated by A2ML1's thioester) to achieve inhibition. In contrast to the A2M tetramer which traps proteases in two internal chambers formed by four subunits, in protease-cleaved monomeric A2ML1 disordered regions surround the trapped protease and may prevent substrate access. In native A2ML1, the bait region is threaded through a hydrophobic channel, suggesting that disruption of this arrangement by bait region cleavage triggers the extensive conformational changes that result in protease inhibition. Structural comparisons with complement C3/C4 suggest that the A2M superfamily of proteins share this mechanism for the triggering of conformational change occurring upon proteolytic activation.
    DOI:  https://doi.org/10.1038/s41467-022-30758-x
  49. Sci Signal. 2022 May 31. 15(736): eabg5216
    Kinase-deficient BTK mutants confer ibrutinib resistance through activation of the kinase HCK.
    Kamaldeep Dhami, Anirban Chakraborty, Tarikere L Gururaja, Leo W-K Cheung, Chaohong Sun, Felix DeAnda, XiaoDong Huang.
      The Bruton's tyrosine kinase (BTK) inhibitor ibrutinib irreversibly binds BTK at Cys481, inhibiting its kinase activity and thus blocking transduction of B cell receptor (BCR) signaling. Although ibrutinib is durably effective in patients with B cell malignancies, many patients still develop ibrutinib-resistant disease. Resistance can arise because of mutations at the ibrutinib-binding site in BTK. Here, we characterized the mechanism by which two BTK mutations, C481F and C481Y, may lead to ibrutinib resistance. Both mutants lacked detectable kinase activity in in vitro kinase assays. Structural modeling suggested that bulky Phe and Tyr side chains at position 481 sterically hinder access to the ATP-binding pocket in BTK, contributing to loss of kinase activity. Nonetheless, BCR signaling still propagated through BTK C481F and C481Y mutants to downstream effectors, the phospholipase PLCγ2 and the transcription factor NF-κB. This maintenance of BCR signaling was partially achieved through the physical recruitment and kinase-independent activation of hematopoietic cell kinase (HCK). Upon BCR activation, BTK C481F or C481Y was phosphorylated by Src family kinases at Tyr551, which then bound to the SH2 domain of HCK. Modeling suggested that this binding disrupted an intramolecular autoinhibitory interaction in HCK. Activated HCK subsequently phosphorylated PLCγ2, which propagated BCR signaling and promoted clonogenic cell proliferation. This kinase-independent mechanism could inform therapeutic approaches to CLL bearing either the C481F or C481Y BTK mutants.
    DOI:  https://doi.org/10.1126/scisignal.abg5216
  50. Curr Opin Genet Dev. 2022 May 26. pii: S0959-437X(22)00030-2. [Epub ahead of print]75 101921
    Cell identity conversion in liver regeneration after injury.
    Qiang He, Lei Cui, Xiang Yuan, Mengyao Wang, Lijian Hui.
      Cell identity conversion in liver injury is the process that mature cells, specifically hepatocytes or cholangiocytes, convert into cells with other identities, which is found to play a pivotal role in liver regeneration. A better characterization of cell identity conversion will not only facilitate the understanding of liver tissue repair but also the development of novel regenerative therapies. In this review, we discuss the latest advances in cell identity conversion during liver regeneration, including conversions between hepatocytes and cholangiocytes and hepatocyte reprogramming to liver progenitor-like cells. To develop a unified description of cellular states in injury-related liver regeneration, we further propose the quantitative approach to explore cell identity conversion based on the Waddington's landscape.
    DOI:  https://doi.org/10.1016/j.gde.2022.101921
  51. Nat Commun. 2022 May 30. 13(1): 2995
    Long-term hepatitis B virus infection of rhesus macaques requires suppression of host immunity.
    Sreya Biswas, Lauren N Rust, Jochen M Wettengel, Sofiya Yusova, Miranda Fischer, Julien N Carson, Josie Johnson, Lei Wei, Trason Thode, Mohan R Kaadige, Sunil Sharma, Majd Agbaria, Benjamin N Bimber, Thomas Tu, Ulrike Protzer, Alexander Ploss, Jeremy V Smedley, Gershon Golomb, Jonah B Sacha, Benjamin J Burwitz.
      Hepatitis B virus has infected a third of the world's population, and 296 million people are living with chronic infection. Chronic infection leads to progressive liver disease, including hepatocellular carcinoma and liver failure, and there remains no reliable curative therapy. These gaps in our understanding are due, in large part, to a paucity of animal models of HBV infection. Here, we show that rhesus macaques regularly clear acute HBV infection, similar to adult humans, but can develop long-term infection if immunosuppressed. Similar to patients, we longitudinally detected HBV DNA, HBV surface antigen, and HBV e antigen in the serum of experimentally infected animals. In addition, we discovered hallmarks of HBV infection in the liver, including RNA transcription, HBV core and HBV surface antigen translation, and covalently closed circular DNA biogenesis. This pre-clinical animal model will serve to accelerate emerging HBV curative therapies into the clinic.
    DOI:  https://doi.org/10.1038/s41467-022-30593-0
  52. J Clin Invest. 2022 Jun 02. pii: e160852. [Epub ahead of print]
    Purine nucleoside phosphorylase enables dual metabolic checkpoints that prevent T cell immunodeficiency and TLR7-dependent autoimmunity.
    Evan R Abt, Khalid Rashid, Thuc M Le, Suwen Li, Hailey R Lee, Vincent Lok, Luyi Li, Amanda L Creech, Amanda N Labora, Hanna K Mandl, Alex K Lam, Arthur Cho, Valerie Rezek, Nanping Wu, Gabriel Abril-Rodriguez, Ethan W Rosser, Steven D Mittelman, Willy Hugo, Thomas Mehrling, Shanta Bantia, Antoni Ribas, Timothy R Donahue, Gay M Crooks, Ting-Ting Wu, Caius G Radu.
      Purine nucleoside phosphorylase (PNP) enables the breakdown and recycling of guanine nucleosides. PNP insufficiency in humans is paradoxically associated with both immunodeficiency and autoimmunity, but the mechanistic basis for these outcomes is incompletely understood. Here we identify two immune lineage-dependent consequences of PNP inactivation dictated by distinct gene interactions. During T cell development, PNP inactivation is synthetically lethal with down-regulation of the dNTP triphosphohydrolase SAMHD1. This interaction requires deoxycytidine kinase activity and is antagonized by microenvironmental deoxycytidine. In B lymphocytes and macrophages, PNP regulates Toll like receptor 7 signaling by controlling the levels of its (deoxy)guanosine nucleoside ligands. Overriding this regulatory mechanism promotes germinal center formation in the absence of exogenous antigen and accelerates disease in a mouse model of autoimmunity. This work reveals that one purine metabolism gene protects against immunodeficiency and autoimmunity via independent mechanisms operating in distinct immune lineages and identifies PNP as a novel metabolic immune checkpoint.
    Keywords:  Autoimmune diseases; Immunology; Immunotherapy; Metabolism; T cell development
    DOI:  https://doi.org/10.1172/JCI160852
  53. Nat Chem Biol. 2022 Jun 02.
    A fungal dioxygenase CcTet serves as a eukaryotic 6mA demethylase on duplex DNA.
    Yajuan Mu, Lin Zhang, Jingyan Hu, Jiashen Zhou, Hou-Wen Lin, Chuan He, Hong-Zhuan Chen, Liang Zhang.
      N6-methyladenosine (6mA) is a DNA modification that has recently been found to play regulatory roles during mammalian early embryo development and mitochondrial transcription. We found that a dioxygenase CcTet from the fungus Coprinopsis cinerea is also a dsDNA 6mA demethylase. It oxidizes 6mA to the intermediate N6-hydroxymethyladenosine (6hmA) with robust activity of 6mA-containing duplex DNA (dsDNA) as well as isolated genomics DNA. Structural characterization revealed that CcTet utilizes three flexible loop regions and two key residues-D337 and G331-in the active pocket to preferentially recognize substrates on dsDNA. A CcTet D337F mutant protein retained the catalytic activity on 6mA but lost activity on 5-methylcytosine. Our findings uncovered a 6mA demethylase that works on dsDNA, suggesting potential 6mA demethylation in fungi and elucidating 6mA recognition and the catalytic mechanism of CcTet. The CcTet D337F mutant protein also provides a chemical biology tool for future functional manipulation of DNA 6mA in vivo.
    DOI:  https://doi.org/10.1038/s41589-022-01041-3
  54. Cell Mol Life Sci. 2022 May 30. 79(6): 325
    Neuroendocrinal and molecular basis of flight performance in locusts.
    Li Hou, Siyuan Guo, Ding Ding, Baozhen Du, Xianhui Wang.
      Insect flight is a complex physiological process that involves sensory and neuroendocrinal control, efficient energy metabolism, rhythmic muscle contraction, and coordinated wing movement. As a classical study model for insect flight, locusts have attracted much attention from physiologists, behaviorists, and neuroendocrinologists over the past decades. In earlier research, scientists made extensive efforts to explore the hormone regulation of metabolism related to locust flight; however, this work was hindered by the absence of molecular and genetic tools. Recently, the rapid development of molecular and genetic tools as well as multi-omics has greatly advanced our understanding of the metabolic, molecular, and neuroendocrinal basis of long-term flight in locusts. Novel neural and molecular factors modulating locust flight and their regulatory mechanisms have been explored. Moreover, the molecular mechanisms underlying phase-dependent differences in locust flight have also been revealed. Here, we provide a systematic review of locust flight physiology, with emphasis on recent advances in the neuroendocrinal, genetic, and molecular basis. Future research directions and potential challenges are also addressed.
    Keywords:  Aging; Energy metabolism; Flight physiology; Hormone; Neuropeptide; Phase-related flight traits
    DOI:  https://doi.org/10.1007/s00018-022-04344-9
  55. Cell Rep. 2022 May 31. pii: S2211-1247(22)00655-6. [Epub ahead of print]39(9): 110880
    LL-37 transports immunoreactive cGAMP to activate STING signaling and enhance interferon-mediated host antiviral immunity.
    Xubiao Wei, Lulu Zhang, Yinlong Yang, Yanfei Hou, Yifang Xu, Zhimeng Wang, Huili Su, Fangping Han, Jing Han, Peiyuan Liu, Shuiqing Hu, Matthew D Koci, Xuxu Sun, Conggang Zhang.
      Cyclic 2',3'-GMP-AMP (cGAMP) binds to and activates stimulator of interferon genes (STING), which then induces interferons to drive immune responses against tumors and pathogens. Exogenous cGAMP produced by infected and malignant cells and synthetic cGAMP used in immunotherapy must traverse the cell membrane to activate STING in target cells. However, as an anionic hydrophilic molecule, cGAMP is not inherently membrane permeable. Here, we show that LL-37, a human host defense peptide, can function as a transporter of cGAMP. LL-37 specifically binds cGAMP and efficiently delivers cGAMP into target cells. cGAMP transferred by LL-37 activates robust interferon responses and host antiviral immunity in a STING-dependent manner. Furthermore, we report that LL-37 inducers vitamin D3 and sodium butyrate promote host immunity by enhancing endogenous LL-37 expression and its mediated cGAMP immune response. Collectively, our data uncover an essential role of LL-37 in innate immune activation and suggest new strategies for immunotherapy.
    Keywords:  CP: Immunology; antibacterial peptide; cGAMP; cGAS-STING signaling pathway; innate immunity; type I interferon
    DOI:  https://doi.org/10.1016/j.celrep.2022.110880
  56. Nat Commun. 2022 May 30. 13(1): 3004
    Stochastic expression of invasion genes in Plasmodium falciparum schizonts.
    Jaishree Tripathi, Lei Zhu, Sourav Nayak, Michal Stoklasa, Zbynek Bozdech.
      Genetically identical cells are known to exhibit differential phenotypes in the same environmental conditions. These phenotypic variants are linked to transcriptional stochasticity and have been shown to contribute towards adaptive flexibility of a wide range of unicellular organisms. Here, we investigate transcriptional heterogeneity and stochastic gene expression in Plasmodium falciparum by performing the quasilinear multiple annealing and looping based amplification cycles (MALBAC) based amplification and single cell RNA sequencing of blood stage schizonts. Our data reveals significant transcriptional variations in the schizont stage with a distinct group of highly variable invasion gene transcripts being identified. Moreover, the data reflects several diversification processes including putative developmental "checkpoint"; transcriptomically distinct parasite sub-populations and transcriptional switches in variable gene families (var, rifin, phist). Most of these features of transcriptional variability are preserved in isogenic parasite cell populations (albeit with a lesser amplitude) suggesting a role of epigenetic factors in cell-to-cell transcriptional variations in human malaria parasites. Lastly, we apply quantitative RT-PCR and RNA-FISH approach and confirm stochastic expression of key invasion genes, such as, msp1, msp3, msp7, eba181 and ama1 which represent prime candidates for invasion-blocking vaccines.
    DOI:  https://doi.org/10.1038/s41467-022-30605-z
  57. Cell Rep. 2022 May 31. pii: S2211-1247(22)00671-4. [Epub ahead of print]39(9): 110896
    CD4 T cells are rapidly depleted from tuberculosis granulomas following acute SIV co-infection.
    Tuberculosis Imaging Program
      HIV/Mycobacterium tuberculosis (Mtb) co-infected individuals have an increased risk of tuberculosis prior to loss of peripheral CD4 T cells, raising the possibility that HIV co-infection leads to CD4 T cell depletion in lung tissue before it is evident in blood. Here, we use rhesus macaques to study the early effects of simian immunodeficiency virus (SIV) co-infection on pulmonary granulomas. Two weeks after SIV inoculation of Mtb-infected macaques, Mtb-specific CD4 T cells are dramatically depleted from granulomas, before CD4 T cell loss in blood, airways, and lymph nodes, or increases in bacterial loads or radiographic evidence of disease. Spatially, CD4 T cells are preferentially depleted from the granuloma core and cuff relative to B cell-rich regions. Moreover, live imaging of granuloma explants show that intralesional CD4 T cell motility is reduced after SIV co-infection. Thus, granuloma CD4 T cells may be decimated before many co-infected individuals experience the first symptoms of acute HIV infection.
    Keywords:  CD4 T cells; CP: Immunology; CP: Microbiology; SIV; granulomas; live imaging; tuberculosis
    DOI:  https://doi.org/10.1016/j.celrep.2022.110896
  58. Front Immunol. 2022 ;13 881656
    Helix-Loop-Helix Proteins in Adaptive Immune Development.
    Megan Aubrey, Zachary J Warburg, Cornelis Murre.
      The E/ID protein axis is instrumental for defining the developmental progression and functions of hematopoietic cells. The E proteins are dimeric transcription factors that activate gene expression programs and coordinate changes in chromatin organization. Id proteins are antagonists of E protein activity. Relative levels of E/Id proteins are modulated throughout hematopoietic development to enable the progression of hematopoietic stem cells into multiple adaptive and innate immune lineages including natural killer cells, B cells and T cells. In early progenitors, the E proteins promote commitment to the T and B cell lineages by orchestrating lineage specific programs of gene expression and regulating VDJ recombination of antigen receptor loci. In mature B cells, the E/Id protein axis functions to promote class switch recombination and somatic hypermutation. E protein activity further regulates differentiation into distinct CD4+ and CD8+ T cells subsets and instructs mature T cell immune responses. In this review, we discuss how the E/Id proteins define the adaptive immune system lineages, focusing on their role in directing developmental gene programs.
    Keywords:  B cell development; E proteins; HLH; Id proteins; T cell development; VDJ recombination; hematopoiesis; lymphopoiesis
    DOI:  https://doi.org/10.3389/fimmu.2022.881656
  59. Sci Rep. 2022 May 31. 12(1): 9033
    Islet cell replacement and transplantation immunology in a mouse strain with inducible diabetes.
    Preksha Bhagchandani, Charles A Chang, Weichen Zhao, Luiza Ghila, Pedro L Herrera, Simona Chera, Seung K Kim.
      Improved models of experimental diabetes are needed to develop cell therapies for diabetes. Here, we introduce the B6 RIP-DTR mouse, a model of experimental diabetes in fully immunocompetent animals. These inbred mice harbor the H2b major histocompatibility complex (MHC), selectively express high affinity human diphtheria toxin receptor (DTR) in islet β-cells, and are homozygous for the Ptprca (CD45.1) allele rather than wild-type Ptprcb (CD45.2). 100% of B6 RIP-DTR mice rapidly became diabetic after a single dose of diphtheria toxin, and this was reversed indefinitely after transplantation with islets from congenic C57BL/6 mice. By contrast, MHC-mismatched islets were rapidly rejected, and this allotransplant response was readily monitored via blood glucose and graft histology. In peripheral blood of B6 RIP-DTR with mixed hematopoietic chimerism, CD45.2 BALB/c donor blood immune cells were readily distinguished from host CD45.1 cells by flow cytometry. Reliable diabetes induction and other properties in B6 RIP-DTR mice provide an important new tool to advance transplant-based studies of islet replacement and immunomodulation to treat diabetes.
    DOI:  https://doi.org/10.1038/s41598-022-13087-3
  60. Nat Commun. 2022 Jun 01. 13(1): 3040
    De novo biosynthesis of rubusoside and rebaudiosides in engineered yeasts.
    Yameng Xu, Xinglong Wang, Chenyang Zhang, Xuan Zhou, Xianhao Xu, Luyao Han, Xueqin Lv, Yanfeng Liu, Song Liu, Jianghua Li, Guocheng Du, Jian Chen, Rodrigo Ledesma-Amaro, Long Liu.
      High-sugar diet causes health problems, many of which can be addressed with the use of sugar substitutes. Rubusoside and rebaudiosides are interesting molecules, considered the next generation of sugar substitutes due to their low-calorie, superior sweetness and organoleptic properties. However, their low abundance in nature makes the traditional plant extraction process neither economical nor environmental-friendly. Here we engineer baker's yeast Saccharomyces cerevisiae as a chassis for the de novo production of rubusoside and rebaudiosides. In this process, we identify multiple issues that limit the production, including rate-liming steps, product stress on cellular fitness and unbalanced metabolic networks. We carry out a systematic engineering strategy to solve these issues, which produces rubusoside and rebaudiosides at titers of 1368.6 mg/L and 132.7 mg/L, respectively. The rubusoside chassis strain here constructed paves the way towards a sustainable, large-scale fermentation-based manufacturing of diverse rebaudiosides.
    DOI:  https://doi.org/10.1038/s41467-022-30826-2
  61. Nature. 2022 Jun 01.
    Spatiotemporal dynamics of noradrenaline during learned behaviour.
    Vincent Breton-Provencher, Gabrielle T Drummond, Jiesi Feng, Yulong Li, Mriganka Sur.
      Noradrenaline released from the locus coeruleus (LC) is a ubiquitous neuromodulator1-4 that has been linked to multiple functions including arousal5-8, action and sensory gain9-11, and learning12-16. Whether and how activation of noradrenaline-expressing neurons in the LC (LC-NA) facilitates different components of specific behaviours is unknown. Here we show that LC-NA activity displays distinct spatiotemporal dynamics to enable two functions during learned behaviour: facilitating task execution and encoding reinforcement to improve performance accuracy. To examine these functions, we used a behavioural task in mice with graded auditory stimulus detection and task performance. Optogenetic inactivation of the LC demonstrated that LC-NA activity was causal for both task execution and optimization. Targeted recordings of LC-NA neurons using photo-tagging, two-photon micro-endoscopy and two-photon output monitoring showed that transient LC-NA activation preceded behavioural execution and followed reinforcement. These two components of phasic activity were heterogeneously represented in LC-NA cortical outputs, such that the behavioural response signal was higher in the motor cortex and facilitated task execution, whereas the negative reinforcement signal was widely distributed among cortical regions and improved response sensitivity on the subsequent trial. Modular targeting of LC outputs thus enables diverse functions, whereby some noradrenaline signals are segregated among targets, whereas others are broadly distributed.
    DOI:  https://doi.org/10.1038/s41586-022-04782-2
  62. Nat Struct Mol Biol. 2022 Jun 02.
    Quaternary structure independent folding of voltage-gated ion channel pore domain subunits.
    Cristina Arrigoni, Marco Lolicato, David Shaya, Ahmed Rohaim, Felix Findeisen, Lam-Kiu Fong, Claire M Colleran, Pawel Dominik, Sangwoo S Kim, Jonathan P Schuermann, William F DeGrado, Michael Grabe, Anthony A Kossiakoff, Daniel L Minor.
      Every voltage-gated ion channel (VGIC) has a pore domain (PD) made from four subunits, each comprising an antiparallel transmembrane helix pair bridged by a loop. The extent to which PD subunit structure requires quaternary interactions is unclear. Here, we present crystal structures of a set of bacterial voltage-gated sodium channel (BacNaV) 'pore only' proteins that reveal a surprising collection of non-canonical quaternary arrangements in which the PD tertiary structure is maintained. This context-independent structural robustness, supported by molecular dynamics simulations, indicates that VGIC-PD tertiary structure is independent of quaternary interactions. This fold occurs throughout the VGIC superfamily and in diverse transmembrane and soluble proteins. Strikingly, characterization of PD subunit-binding Fabs indicates that non-canonical quaternary PD conformations can occur in full-length VGICs. Together, our data demonstrate that the VGIC-PD is an autonomously folded unit. This property has implications for VGIC biogenesis, understanding functional states, de novo channel design, and VGIC structural origins.
    DOI:  https://doi.org/10.1038/s41594-022-00775-x
  63. Nature. 2022 Jun 01.
    Molecularly defined circuits for cardiovascular and cardiopulmonary control.
    Avin Veerakumar, Andrea R Yung, Yin Liu, Mark A Krasnow.
      The sympathetic and parasympathetic nervous systems regulate the activities of internal organs1, but the molecular and functional diversity of their constituent neurons and circuits remains largely unknown. Here we use retrograde neuronal tracing, single-cell RNA sequencing, optogenetics and physiological experiments to dissect the cardiac parasympathetic control circuit in mice. We show that cardiac-innervating neurons in the brainstem nucleus ambiguus (Amb) are comprised of two molecularly, anatomically and functionally distinct subtypes. The first, which we call ambiguus cardiovascular (ACV) neurons (approximately 35 neurons per Amb), define the classical cardiac parasympathetic circuit. They selectively innervate a subset of cardiac parasympathetic ganglion neurons and mediate the baroreceptor reflex, slowing heart rate and atrioventricular node conduction in response to increased blood pressure. The other, ambiguus cardiopulmonary (ACP) neurons (approximately 15 neurons per Amb) innervate cardiac ganglion neurons intermingled with and functionally indistinguishable from those innervated by ACV neurons. ACP neurons also innervate most or all lung parasympathetic ganglion neurons-clonal labelling shows that individual ACP neurons innervate both organs. ACP neurons mediate the dive reflex, the simultaneous bradycardia and bronchoconstriction that follows water immersion. Thus, parasympathetic control of the heart is organized into two parallel circuits, one that selectively controls cardiac function (ACV circuit) and another that coordinates cardiac and pulmonary function (ACP circuit). This new understanding of cardiac control has implications for treating cardiac and pulmonary diseases and for elucidating the control and coordination circuits of other organs.
    DOI:  https://doi.org/10.1038/s41586-022-04760-8
  64. J Biol Chem. 2022 May 25. pii: S0021-9258(22)00517-8. [Epub ahead of print] 102077
    Redox regulation by TXNRD3 during epididymal maturation underlies capacitation-associated mitochondrial activity and sperm motility in mice.
    Huafeng Wang, Qianhui Dou, Kyung Jo Jeong, Jungmin Choi, Vadim N Gladyshev, Jean-Ju Chung.
      During epididymal transit, redox remodeling protects mammalian spermatozoa, preparing them for survival in the subsequent journey to fertilization. However, molecular mechanisms of redox regulation in sperm development and maturation remain largely elusive. In this study, we report that TXNRD3, a thioredoxin reductase family member particularly abundant in elongating spermatids at the site of mitochondrial sheath formation, regulates redox homeostasis to support male fertility. Using Txnrd3-/- mice, our biochemical, ultrastructural, and live cell imaging analyses revealed impairments in sperm morphology and motility under conditions of TXNRD3 deficiency. We find that mitochondria develop more defined cristae during capacitation in wild type sperm. Furthermore, we show that absence of TXNRD3 alters thiol redox status in both the head and tail during sperm maturation and capacitation, resulting in defective mitochondrial ultrastructure and activity under capacitating conditions. These findings provide insights into molecular mechanisms of redox homeostasis and bioenergetics during sperm maturation, capacitation, and fertilization.
    Keywords:  Redox homeostasis; TXNRD3; Thioredoxin Glutathione Reductase; Ultrastructure; male fertility; mitochondrial function
    DOI:  https://doi.org/10.1016/j.jbc.2022.102077
  65. Diabetes. 2022 Jun 03. pii: dbi220003. [Epub ahead of print]
    Regulation of Hepatic Lipid and Glucose Metabolism by INSP3R1.
    Rachel J Perry.
      With the rising epidemics of obesity and nonalcoholic fatty liver disease (NAFLD) and its downstream consequences including steatohepatitis, cirrhosis, and type 2 diabetes in the U.S. and worldwide, new therapeutic approaches are urgently needed to treat these devastating conditions. Glucagon, known for a century to be a glucose-raising hormone and clearly demonstrated to contribute to fasting and postprandial hyperglycemia in both type 1 and type 2 diabetes, represents an unlikely target to improve health in those with metabolic syndrome. However, recent work from our group and others' identifies an unexpected role for glucagon as a potential means of treating NAFLD, improving insulin sensitivity, and improving the lipid profile. We propose a unifying, calcium-dependent mechanism for glucagon's effects both to stimulate hepatic gluconeogenesis and to enhance hepatic mitochondrial oxidation: signaling through the inositol 1,4,5-trisphosphate receptor type 1 (INSP3R1), glucagon activates phospholipase C (PKC)/protein kinase A (PKA) signaling to enhance adipose triglyceride lipase (ATGL)-dependent intrahepatic lipolysis and, in turn, increase cytosolic gluconeogenesis by allosteric activation of pyruvate carboxylase. Simultaneously in the mitochondria, calcium transferred through mitochondria-associated membranes activates several dehydrogenases in the tricarboxylic acid cycle, correlated with an increase in mitochondrial energy expenditure and reduction in ectopic lipid. This model suggests that short-term, cyclic treatment with glucagon or other INSP3R1 antagonists could hold promise as a means to reset lipid homeostasis in patients with NAFLD.
    DOI:  https://doi.org/10.2337/dbi22-0003
  66. Nature. 2022 Jun 01.
    Combination anti-HIV antibodies provide sustained virological suppression.
    Michael C Sneller, Jana Blazkova, J Shawn Justement, Victoria Shi, Brooke D Kennedy, Kathleen Gittens, Jekaterina Tolstenko, Genevieve McCormack, Emily J Whitehead, Rachel F Schneck, Michael A Proschan, Erika Benko, Colin Kovacs, Cihan Oguz, Michael S Seaman, Marina Caskey, Michel C Nussenzweig, Anthony S Fauci, Susan Moir, Tae-Wook Chun.
      Antiretroviral therapy is highly effective in suppressing human immunodeficiency virus (HIV)1. However, eradication of the virus in individuals with HIV has not been possible to date2. Given that HIV suppression requires life-long antiretroviral therapy, predominantly on a daily basis, there is a need to develop clinically effective alternatives that use long-acting antiviral agents to inhibit viral replication3. Here we report the results of a two-component clinical trial involving the passive transfer of two HIV-specific broadly neutralizing monoclonal antibodies, 3BNC117 and 10-1074. The first component was a randomized, double-blind, placebo-controlled trial that enrolled participants who initiated antiretroviral therapy during the acute/early phase of HIV infection. The second component was an open-label single-arm trial that enrolled individuals with viraemic control who were naive to antiretroviral therapy. Up to 8 infusions of 3BNC117 and 10-1074, administered over a period of 24 weeks, were well tolerated without any serious adverse events related to the infusions. Compared with the placebo, the combination broadly neutralizing monoclonal antibodies maintained complete suppression of plasma viraemia (for up to 43 weeks) after analytical treatment interruption, provided that no antibody-resistant HIV was detected at the baseline in the study participants. Similarly, potent HIV suppression was seen in the antiretroviral-therapy-naive study participants with viraemia carrying sensitive virus at the baseline. Our data demonstrate that combination therapy with broadly neutralizing monoclonal antibodies can provide long-term virological suppression without antiretroviral therapy in individuals with HIV, and our experience offers guidance for future clinical trials involving next-generation antibodies with long half-lives.
    DOI:  https://doi.org/10.1038/s41586-022-04797-9
  67. Cell Rep. 2022 May 31. pii: S2211-1247(22)00646-5. [Epub ahead of print]39(9): 110871
    Disrupted control of origin activation compromises genome integrity upon destabilization of Polε and dysfunction of the TRP53-CDKN1A/P21 axis.
    Valerie Borel, Stefan Boeing, Niek Van Wietmarschen, Sriram Sridharan, Bethany Rebekah Hill, Luigi Ombrato, Jimena Perez-Lloret, Deb Jackson, Robert Goldstone, Simon J Boulton, Andre Nussenzweig, Roberto Bellelli.
      The maintenance of genome stability relies on coordinated control of origin activation and replication fork progression. How the interplay between these processes influences human genetic disease and cancer remains incompletely characterized. Here we show that mouse cells featuring Polε instability exhibit impaired genome-wide activation of DNA replication origins, in an origin-location-independent manner. Strikingly, Trp53 ablation in primary Polε hypomorphic cells increased Polε levels and origin activation and reduced DNA damage in a transcription-dependent manner. Transcriptome analysis of primary Trp53 knockout cells revealed that the TRP53-CDKN1A/P21 axis maintains appropriate levels of replication factors and CDK activity during unchallenged S phase. Loss of this control mechanism deregulates origin activation and perturbs genome-wide replication fork progression. Thus, while our data support an impaired origin activation model for genetic diseases affecting CMG formation, we propose that loss of the TRP53-CDKN1A/P21 tumor suppressor axis induces inappropriate origin activation and deregulates genome-wide fork progression.
    Keywords:  CDKN1A/P21; CP: Molecular biology; DNA replication; Polε; TRP53; genome stability
    DOI:  https://doi.org/10.1016/j.celrep.2022.110871
  68. Front Cell Dev Biol. 2022 ;10 871326
    Lung Cancer Induces NK Cell Contractility and Cytotoxicity Through Transcription Factor Nuclear Localization.
    Darren Chen Pei Wong, E Hui Clarissa Lee, Junzhi Er, Ivan Yow, Ricky Abdi Gunawan Koean, Owen Ang, Jingwei Xiao, Boon Chuan Low, Jeak Ling Ding.
      Actomyosin-mediated cellular contractility is highly conserved for mechanotransduction and signalling. While this phenomenon has been observed in adherent cell models, whether/how contractile forces regulate the function of suspension cells like natural killer (NK) cells during cancer surveillance, is unknown. Here, we demonstrated in coculture settings that the evolutionarily conserved NK cell transcription factor, Eomes, undergoes nuclear shuttling during lung cancer cell surveillance. Biophysical and biochemical analyses revealed mechanistic enhancement of NK cell actomyosin-mediated contractility, which is associated with nuclear flattening, thus enabling nuclear entry of Eomes associated with enhanced NK cytotoxicity. We found that NK cells responded to the presumed immunosuppressive TGFβ in the NK-lung cancer coculture medium to sustain its intracellular contractility through myosin light chain phosphorylation, thereby promoting Eomes nuclear localization. Therefore, our results demonstrate that lung cancer cells provoke NK cell contractility as an early phase activation mechanism and that Eomes is a plausible mechano-responsive protein for increased NK cytotoxicity. There is scope for strategic application of actomyosin-mediated contractility modulating drugs ex vivo, to reinvigorate NK cells prior to adoptive cancer immunotherapy in vivo (177 words).
    Keywords:  Eomesdermin (Eomes); NK-cancer cell interaction; TGFβ disparate role; actin cytoskeleton; cellular contractility of natural killer (NK) cell; mechanotransduction in NK cells; non-small cell lung cancer (NSCLC, invasive and non-invasive subtypes); transcription factor shuttling
    DOI:  https://doi.org/10.3389/fcell.2022.871326
  69. Nat Commun. 2022 Jun 01. 13(1): 3051
    POWR1 is a domestication gene pleiotropically regulating seed quality and yield in soybean.
    Wolfgang Goettel, Hengyou Zhang, Ying Li, Zhenzhen Qiao, He Jiang, Dianyun Hou, Qijian Song, Vincent R Pantalone, Bao-Hua Song, Deyue Yu, Yong-Qiang Charles An.
      Seed protein, oil content and yield are highly correlated agronomically important traits that essentially account for the economic value of soybean. The underlying molecular mechanisms and selection of these correlated seed traits during soybean domestication are, however, less known. Here, we demonstrate that a CCT gene, POWR1, underlies a large-effect protein/oil QTL. A causative TE insertion truncates its CCT domain and substantially increases seed oil content, weight, and yield while decreasing protein content. POWR1 pleiotropically controls these traits likely through regulating seed nutrient transport and lipid metabolism genes. POWR1 is also a domestication gene. We hypothesize that the TE insertion allele is exclusively fixed in cultivated soybean due to selection for larger seeds during domestication, which significantly contributes to shaping soybean with increased yield/seed weight/oil but reduced protein content. This study provides insights into soybean domestication and is significant in improving seed quality and yield in soybean and other crop species.
    DOI:  https://doi.org/10.1038/s41467-022-30314-7
  70. J Exp Med. 2022 Jul 04. pii: e20220017. [Epub ahead of print]219(7):
    Microbiota and adipocyte mitochondrial damage in type 2 diabetes are linked by Mmp12+ macrophages.
    Zhipeng Li, Manoj Gurung, Richard R Rodrigues, Jyothi Padiadpu, Nolan K Newman, Nathan P Manes, Jacob W Pederson, Renee L Greer, Stephany Vasquez-Perez, Hyekyoung You, Kaito A Hioki, Zoe Moulton, Anna Fel, Dominic De Nardo, Amiran K Dzutsev, Aleksandra Nita-Lazar, Giorgio Trinchieri, Natalia Shulzhenko, Andrey Morgun.
      Microbiota contribute to the induction of type 2 diabetes by high-fat/high-sugar (HFHS) diet, but which organs/pathways are impacted by microbiota remain unknown. Using multiorgan network and transkingdom analyses, we found that microbiota-dependent impairment of OXPHOS/mitochondria in white adipose tissue (WAT) plays a primary role in regulating systemic glucose metabolism. The follow-up analysis established that Mmp12+ macrophages link microbiota-dependent inflammation and OXPHOS damage in WAT. Moreover, the molecular signature of Mmp12+ macrophages in WAT was associated with insulin resistance in obese patients. Next, we tested the functional effects of MMP12 and found that Mmp12 genetic deficiency or MMP12 inhibition improved glucose metabolism in conventional, but not in germ-free mice. MMP12 treatment induced insulin resistance in adipocytes. TLR2-ligands present in Oscillibacter valericigenes bacteria, which are expanded by HFHS, induce Mmp12 in WAT macrophages in a MYD88-ATF3-dependent manner. Thus, HFHS induces Mmp12+ macrophages and MMP12, representing a microbiota-dependent bridge between inflammation and mitochondrial damage in WAT and causing insulin resistance.
    DOI:  https://doi.org/10.1084/jem.20220017
  71. Cell Rep. 2022 May 31. pii: S2211-1247(22)00660-X. [Epub ahead of print]39(9): 110885
    Intrinsic antiviral immunity of barrier cells revealed by an iPSC-derived blood-brain barrier cellular model.
    Yichen Cheng, Angelica Medina, Zhenlan Yao, Mausumi Basu, Janhavi P Natekar, Jianshe Lang, Egan Sanchez, Mezindia B Nkembo, Chongchong Xu, Xuyu Qian, Phuong T T Nguyen, Zhexing Wen, Hongjun Song, Guo-Li Ming, Mukesh Kumar, Margo A Brinton, Melody M H Li, Hengli Tang.
      Physiological blood-tissue barriers play a critical role in separating the circulation from immune-privileged sites and denying access to blood-borne viruses. The mechanism of virus restriction by these barriers is poorly understood. We utilize induced pluripotent stem cell (iPSC)-derived human brain microvascular endothelial cells (iBMECs) to study virus-blood-brain barrier (BBB) interactions. These iPSC-derived cells faithfully recapitulate a striking difference in in vivo neuroinvasion by two alphavirus isolates and are selectively permissive to neurotropic flaviviruses. A model of cocultured iBMECs and astrocytes exhibits high transendothelial electrical resistance and blocks non-neurotropic flaviviruses from getting across the barrier. We find that iBMECs constitutively express an interferon-induced gene, IFITM1, which preferentially restricts the replication of non-neurotropic flaviviruses. Barrier cells from blood-testis and blood-retinal barriers also constitutively express IFITMs that contribute to the viral resistance. Our application of a renewable human iPSC-based model for studying virus-BBB interactions reveals that intrinsic immunity at the barriers contributes to virus exclusion.
    Keywords:  CP: Immunology; CP: Neuroscience; IFITM; Zika virus; blood-brain barrier; blood-retinal barrier; blood-testis barrier; brain microvascular endothelial cell; dengue virus; flavivirus; intrinsic expression; virus-host interactions
    DOI:  https://doi.org/10.1016/j.celrep.2022.110885
  72. Cancer Discov. 2022 Jun 02. 12(6): 1410-1412
    Epigenetic Regulation to Enhance Graft-versus-Leukemia Activity.
    Natalie Köhler, Robert Zeiser.
       SUMMARY: Accumulating evidence supports that loss of HLA expression contributes to relapse after allogeneic hematopoietic cell transplantation (allo-HCT), but the mechanisms behind this evasion strategy are unclear. The groups of Luca Vago and Raffaella Di Micco identified the polycomb repressive complex 2 (PRC2) as a key epigenetic driver of immune escape after allo-HCT by reducing the chromatin accessibility of HLA class II molecules, which could be targeted by pharmacologic inhibition of PRC2 subunits. See related article by Gambacorta et al., p. 1449 (10).
    DOI:  https://doi.org/10.1158/2159-8290.CD-22-0329
  73. Nat Commun. 2022 Jun 01. 13(1): 3061
    The genomic landscape of cholangiocarcinoma reveals the disruption of post-transcriptional modifiers.
    Yaodong Zhang, Zijian Ma, Changxian Li, Cheng Wang, Wangjie Jiang, Jiang Chang, Sheng Han, Zefa Lu, Zicheng Shao, Yirui Wang, Hongwei Wang, Chenyu Jiao, Dong Wang, Xiaofeng Wu, Hongbing Shen, Xuehao Wang, Zhibin Hu, Xiangcheng Li.
      Molecular variation between geographical populations and subtypes indicate potential genomic heterogeneity and novel genomic features within CCA. Here, we analyze exome-sequencing data of 87 perihilar cholangiocarcinoma (pCCA) and 261 intrahepatic cholangiocarcinoma (iCCA) cases from 3 Asian centers (including 43 pCCAs and 24 iCCAs from our center). iCCA tumours demonstrate a higher tumor mutation burden and copy number alteration burden (CNAB) than pCCA tumours, and high CNAB indicates a poorer pCCA prognosis. We identify 12 significantly mutated genes and 5 focal CNA regions, and demonstrate common mutations in post-transcriptional modification-related potential driver genes METTL14 and RBM10 in pCCA tumours. Finally we demonstrate the tumour-suppressive role of METTL14, a major RNA N6-adenosine methyltransferase (m6A), and illustrate that its loss-of-function mutation R298H may act through m6A modification on potential driver gene MACF1. Our results may be valuable for better understanding of how post-transcriptional modification can affect CCA development, and highlight both similarities and differences between pCCA and iCCA.
    DOI:  https://doi.org/10.1038/s41467-022-30708-7
  74. EBioMedicine. 2022 May 27. pii: S2352-3964(22)00260-2. [Epub ahead of print]81 104079
    Common risk variants for epilepsy are enriched in families previously targeted for rare monogenic variant discovery.
    Epi4K Consortium
       BACKGROUND: The epilepsies are highly heritable conditions that commonly follow complex inheritance. While monogenic causes have been identified in rare familial epilepsies, most familial epilepsies remain unsolved. We aimed to determine (1) whether common genetic variation contributes to familial epilepsy risk, and (2) whether that genetic risk is enriched in familial compared with non-familial (sporadic) epilepsies.
    METHODS: Using common variants derived from the largest epilepsy genome-wide association study, we calculated polygenic risk scores (PRS) for patients with familial epilepsy (n = 1,818 from 1,181 families), their unaffected relatives (n = 771), sporadic patients (n = 1,182), and population controls (n = 15,929). We also calculated separate PRS for genetic generalised epilepsy (GGE) and focal epilepsy. Statistical analyses used mixed-effects regression models to account for familial relatedness, sex, and ancestry.
    FINDINGS: Patients with familial epilepsies had higher epilepsy PRS compared to population controls (OR 1·20, padj = 5×10-9), sporadic patients (OR 1·11, padj = 0.008), and their own unaffected relatives (OR 1·12, padj = 0.01). The top 1% of the PRS distribution was enriched 3.8-fold for individuals with familial epilepsy when compared to the lowest decile (padj = 5×10-11). Familial PRS enrichment was consistent across epilepsy type; overall, polygenic risk was greatest for the GGE clinical group. There was no significant PRS difference in familial cases with established rare variant genetic etiologies compared to unsolved familial cases.
    INTERPRETATION: The aggregate effects of common genetic variants, measured as polygenic risk scores, play an important role in explaining why some families develop epilepsy, why specific family members are affected while their relatives are not, and why families manifest specific epilepsy types. Polygenic risk contributes to the complex inheritance of the epilepsies, including in individuals with a known genetic etiology.
    FUNDING: National Health and Medical Research Council of Australia, National Institutes of Health, American Academy of Neurology, Thomas B and Jeannette E Laws McCabe Fund, Mirowski Family Foundation.
    Keywords:  Common genetic variation; Epilepsy genetics; Familial epilepsies; Polygenic risk scores
    DOI:  https://doi.org/10.1016/j.ebiom.2022.104079
  75. Nat Commun. 2022 May 30. 13(1): 2994
    Identification of positive and negative regulators of antiviral RNA interference in Arabidopsis thaliana.
    Si Liu, Meijuan Chen, Ruidong Li, Wan-Xiang Li, Amit Gal-On, Zhenyu Jia, Shou-Wei Ding.
      Virus-host coevolution often drives virus immune escape. However, it remains unknown whether natural variations of plant virus resistance are enriched in genes of RNA interference (RNAi) pathway known to confer essential antiviral defense in plants. Here, we report two genome-wide association study screens to interrogate natural variation among wild-collected Arabidopsis thaliana accessions in quantitative resistance to the endemic cucumber mosaic virus (CMV). We demonstrate that the highest-ranked gene significantly associated with resistance from both screens acts to regulate antiviral RNAi in ecotype Columbia-0. One gene, corresponding to Reduced Dormancy 5 (RDO5), enhances resistance by promoting amplification of the virus-derived small interfering RNAs (vsiRNAs). Interestingly, the second gene, designated Antiviral RNAi Regulator 1 (VIR1), dampens antiviral RNAi so its genetic inactivation by CRISPR/Cas9 editing enhances both vsiRNA production and CMV resistance. Our findings identify positive and negative regulators of the antiviral RNAi defense that may play important roles in virus-host coevolution.
    DOI:  https://doi.org/10.1038/s41467-022-30771-0
  76. Nat Cell Biol. 2022 Jun 02.
    A stromal Integrated Stress Response activates perivascular cancer-associated fibroblasts to drive angiogenesis and tumour progression.
    Ioannis I Verginadis, Harris Avgousti, James Monslow, Giorgos Skoufos, Frank Chinga, Kyle Kim, Nektaria Maria Leli, Ilias V Karagounis, Brett I Bell, Anastasia Velalopoulou, Carlo Salas Salinas, Victoria S Wu, Yang Li, Jiangbin Ye, David A Scott, Andrei L Osterman, Arjun Sengupta, Aalim Weljie, Menggui Huang, Duo Zhang, Yi Fan, Enrico Radaelli, John W Tobias, Florian Rambow, Panagiotis Karras, Jean-Christophe Marine, Xiaowei Xu, Artemis G Hatzigeorgiou, Sandra Ryeom, J Alan Diehl, Serge Y Fuchs, Ellen Puré, Constantinos Koumenis.
      Bidirectional signalling between the tumour and stroma shapes tumour aggressiveness and metastasis. ATF4 is a major effector of the Integrated Stress Response, a homeostatic mechanism that couples cell growth and survival to bioenergetic demands. Using conditional knockout ATF4 mice, we show that global, or fibroblast-specific loss of host ATF4, results in deficient vascularization and a pronounced growth delay of syngeneic melanoma and pancreatic tumours. Single-cell transcriptomics of tumours grown in Atf4Δ/Δ mice uncovered a reduction in activation markers in perivascular cancer-associated fibroblasts (CAFs). Atf4Δ/Δ fibroblasts displayed significant defects in collagen biosynthesis and deposition and a reduced ability to support angiogenesis. Mechanistically, ATF4 regulates the expression of the Col1a1 gene and levels of glycine and proline, the major amino acids of collagen. Analyses of human melanoma and pancreatic tumours revealed a strong correlation between ATF4 and collagen levels. Our findings establish stromal ATF4 as a key driver of CAF functionality, malignant progression and metastasis.
    DOI:  https://doi.org/10.1038/s41556-022-00918-8
  77. J Pharmacol Sci. 2022 Jul;pii: S1347-8613(22)00033-0. [Epub ahead of print]149(3): 166-171
    The transcription factor Hhex regulates inflammation-related genes in microglia.
    Risa Sakate, Masahiro Nishiyama, Yu Fukuda, Shiho Kitaoka, Tomoyuki Furuyashiki.
      Microglia have diverse physiological and pathological functions. However, the transcriptional mechanisms remain elusive. Here we sought new transcription factors relevant to microglial functions from the microglial transcriptome of stressed mice and evaluated their roles in primary microglia. TLR2 and TLR4 agonists increased Rel, Atf3, and Cebpb and decreased Hhex in primary microglia as repeated social defeat stress. Although Hhex was not studied in microglia, TLR2 and TLR4 agonists decreased Hhex, and Hhex overexpression attenuated TLR4-increased expression of inflammation-related genes. These findings suggest that Hhex negatively regulates inflammation-related genes in microglia and that TLR2/4 activation reduces Hhex, facilitating TLR4-mediated neuroinflammation.
    Keywords:  Hhex; Inflammation; Microglia
    DOI:  https://doi.org/10.1016/j.jphs.2022.04.006
  78. Nat Chem Biol. 2022 May 30.
    Ribosome stalling during selenoprotein translation exposes a ferroptosis vulnerability.
    Zhipeng Li, Lucas Ferguson, Kirandeep K Deol, Melissa A Roberts, Leslie Magtanong, Joseph M Hendricks, Gergey Alzaem Mousa, Seda Kilinc, Kaitlin Schaefer, James A Wells, Michael C Bassik, Andrei Goga, Scott J Dixon, Nicholas T Ingolia, James A Olzmann.
      The selenoprotein glutathione peroxidase 4 (GPX4) prevents ferroptosis by converting lipid peroxides into nontoxic lipid alcohols. GPX4 has emerged as a promising therapeutic target for cancer treatment, but some cancer cells are resistant to ferroptosis triggered by GPX4 inhibition. Using a chemical-genetic screen, we identify LRP8 (also known as ApoER2) as a ferroptosis resistance factor that is upregulated in cancer. Loss of LRP8 decreases cellular selenium levels and the expression of a subset of selenoproteins. Counter to the canonical hierarchical selenoprotein regulatory program, GPX4 levels are strongly reduced due to impaired translation. Mechanistically, low selenium levels result in ribosome stalling at the inefficiently decoded GPX4 selenocysteine UGA codon, leading to ribosome collisions, early translation termination and proteasomal clearance of the N-terminal GPX4 fragment. These findings reveal rewiring of the selenoprotein hierarchy in cancer cells and identify ribosome stalling and collisions during GPX4 translation as ferroptosis vulnerabilities in cancer.
    DOI:  https://doi.org/10.1038/s41589-022-01033-3
  79. Front Immunol. 2022 ;13 880887
    Macrophage Cx43 Is Necessary for Fibroblast Cytosolic Calcium and Lung Fibrosis After Injury.
    Aritra Bhattacharyya, Paola Torre, Preeti Yadav, Kaveh Boostanpour, Tian Y Chen, Tatsuya Tsukui, Dean Sheppard, Rieko Muramatsu, Robert I Seed, Stephen L Nishimura, James B Jung, Xin-Zi Tang, Christopher D C Allen, Mallar Bhattacharya.
      Macrophages are paracrine signalers that regulate tissular responses to injury through interactions with parenchymal cells. Connexin hemichannels have recently been shown to mediate efflux of ATP by macrophages, with resulting cytosolic calcium responses in adjacent cells. Here we report that lung macrophages with deletion of connexin 43 (MacΔCx43) had decreased ATP efflux into the extracellular space and induced a decreased cytosolic calcium response in co-cultured fibroblasts compared to WT macrophages. Furthermore, MacΔCx43 mice had decreased lung fibrosis after bleomycin-induced injury. Interrogating single cell data for human and mouse, we found that P2rx4 was the most highly expressed ATP receptor and calcium channel in lung fibroblasts and that its expression was increased in the setting of fibrosis. Fibroblast-specific deletion of P2rx4 in mice decreased lung fibrosis and collagen expression in lung fibroblasts in the bleomycin model. Taken together, these studies reveal a Cx43-dependent profibrotic effect of lung macrophages and support development of fibroblast P2rx4 as a therapeutic target for lung fibrosis.
    Keywords:  P2RX4; calcium imaging; fibroblast; lung fibrosis; macrophage
    DOI:  https://doi.org/10.3389/fimmu.2022.880887
  80. Proc Natl Acad Sci U S A. 2022 Jun 07. 119(23): e2121335119
    Human pathogenic RNA viruses establish noncompeting lineages by occupying independent niches.
    Pascal Mutz, Nash D Rochman, Yuri I Wolf, Guilhem Faure, Feng Zhang, Eugene V Koonin.
      SignificanceNumerous pathogenic viruses are endemic in humans and cause a broad variety of diseases, but what is their potential for causing new pandemics? We show that most human pathogenic RNA viruses form multiple, cocirculating lineages with low turnover rates. These lineages appear to be largely noncompeting and occupy distinct epidemiological niches that are not regionally or seasonally defined, and their persistence appears to stem from limited outbreaks in small communities so that only a small fraction of the global susceptible population is infected at any time. However, due to globalization, interaction and competition between lineages might increase, potentially leading to increased diversification and pathogenicity. Thus, endemic viruses appear to merit global attention with respect to the prevention of future pandemics.
    Keywords:  effective population size; globalization; human RNA viruses; influenza; pandemic
    DOI:  https://doi.org/10.1073/pnas.2121335119
  81. J Mol Med (Berl). 2022 Jun;100(6): 963-971
    Leukocyte cytokine responses in adult patients with mitochondrial DNA defects.
    Kalpita R Karan, Caroline Trumpff, Marissa Cross, Kristin M Engelstad, Anna L Marsland, Peter J McGuire, Michio Hirano, Martin Picard.
      Patients with oxidative phosphorylation (OxPhos) defects causing mitochondrial diseases appear particularly vulnerable to infections. Although OxPhos defects modulate cytokine production in vitro and in animal models, little is known about how circulating leukocytes of patients with inherited mitochondrial DNA (mtDNA) defects respond to acute immune challenges. In a small cohort of healthy controls (n = 21) and patients (n = 12) with either the m.3243A > G mutation or single, large-scale mtDNA deletions, we examined (i) cytokine responses (IL-6, TNF-α, IL-1β) in response to acute lipopolysaccharide (LPS) exposure and (ii) sensitivity to the immunosuppressive effects of glucocorticoid signaling (dexamethasone) on cytokine production. In dose-response experiments to determine the half-maximal effective LPS concentration (EC50), relative to controls, leukocytes from patients with mtDNA deletions showed 74-79% lower responses for IL-6 and IL-1β (pIL-6 = 0.031, pIL-1β = 0.009). Moreover, whole blood from patients with mtDNA deletions (pIL-6 = 0.006), but not patients with the m.3243A > G mutation, showed greater sensitivity to the immunosuppressive effects of dexamethasone. Together, these ex vivo data provide preliminary evidence that some systemic OxPhos defects may compromise immune cytokine responses and increase the sensitivity to immune cytokine suppression by glucocorticoids. Further work in larger cohorts is needed to define the nature of immune dysregulation in patients with mitochondrial disease, and their potential implications for disease phenotypes. KEY MESSAGES: Little is known about leukocyte cytokine responses in patients with mitochondrial diseases. Leukocytes of patients with mtDNA deletions show blunted LPS sensitivity and cytokine responses. Leukocytes of patients with mtDNA deletions are more sensitive to glucocorticoid-mediated IL-6 suppression. Work in larger cohorts is needed to delineate potential immune alterations in mitochondrial diseases.
    Keywords:  3243A > G; Cytokine; Glucocorticoid; Inflammation; Inflammation Suppression; Interleukin; Mitochondrial disease; mtDNA deletion
    DOI:  https://doi.org/10.1007/s00109-022-02206-2
  82. Nat Commun. 2022 May 31. 13(1): 3026
    Modular (de)construction of complex bacterial phenotypes by CRISPR/nCas9-assisted, multiplex cytidine base-editing.
    Daniel C Volke, Román A Martino, Ekaterina Kozaeva, Andrea M Smania, Pablo I Nikel.
      CRISPR/Cas technologies constitute a powerful tool for genome engineering, yet their use in non-traditional bacteria depends on host factors or exogenous recombinases, which limits both efficiency and throughput. Here we mitigate these practical constraints by developing a widely-applicable genome engineering toolset for Gram-negative bacteria. The challenge is addressed by tailoring a CRISPR base editor that enables single-nucleotide resolution manipulations (C·G → T·A) with >90% efficiency. Furthermore, incorporating Cas6-mediated processing of guide RNAs in a streamlined protocol for plasmid assembly supports multiplex base editing with >85% efficiency. The toolset is adopted to construct and deconstruct complex phenotypes in the soil bacterium Pseudomonas putida. Single-step engineering of an aromatic-compound production phenotype and multi-step deconstruction of the intricate redox metabolism illustrate the versatility of multiplex base editing afforded by our toolbox. Hence, this approach overcomes typical limitations of previous technologies and empowers engineering programs in Gram-negative bacteria that were out of reach thus far.
    DOI:  https://doi.org/10.1038/s41467-022-30780-z
  83. Proc Natl Acad Sci U S A. 2022 Jun 07. 119(23): e2201794119
    GPR174 signals via Gαs to control a CD86-containing gene expression program in B cells.
    Elise W Wolf, Zachary P Howard, Lihui Duan, Hanson Tam, Ying Xu, Jason G Cyster.
      SignificanceModeling immune responses in vitro is critical for studying many facets of the B cell response. We show that during culture without stimulation, mouse B cells undergo massive changes in gene expression. Many of these changes are promoted by GPR174 signaling via Gαs. GPR174 and Gαs also contribute to reduced B cell viability during culture. We suggest that GPR174 antagonists may be useful to reduce the shift in gene expression and to augment B cell survival during culture. We also provide evidence that ligand engagement of GPR174 can activate this pathway in vivo. Variants in the GPR174 locus have been associated with autoimmune diseases. Our findings provide knowledge for understanding how alterations in GPR174 expression may contribute to disease.
    Keywords:  B lymphocytes; CD86; G-protein–coupled receptor; GPR174; NUR77
    DOI:  https://doi.org/10.1073/pnas.2201794119
  84. Sci Rep. 2022 Jun 01. 12(1): 9142
    Lifelong changes of neurotransmitter receptor expression and debilitation of hippocampal synaptic plasticity following early postnatal blindness.
    Hardy Hagena, Mirko Feldmann, Denise Manahan-Vaughan.
      In the weeks immediately after onset of sensory loss, extensive reorganization of both the cortex and hippocampus occurs. Two fundamental characteristics comprise widespread changes in the relative expression of GABA and glutamate receptors and debilitation of hippocampal synaptic plasticity. Here, we explored whether recovery from adaptive changes in the expression of plasticity-related neurotransmitter receptors and hippocampal synaptic plasticity occurs in the time-period of up to 12 months after onset of sensory loss. We compared receptor expression in CBA/J mice that develop hereditary blindness, with CBA/CaOlaHsd mice that have intact vision and no deficits in other sensory modalities throughout adulthood. GluN1-subunit expression was reduced and the GluN2A:GluN2B ratio was persistently altered in cortex and hippocampus. GABA-receptor expression was decreased and metabotropic glutamate receptor expression was altered. Hippocampal synaptic plasticity was persistently compromised in vivo. But although LTP in blind mice was chronically impaired throughout adulthood, a recovery of the early phase of LTP became apparent when the animals reached 12 months of age. These data show that cortical and hippocampal adaptation to early postnatal blindness progresses into advanced adulthood and is a process that compromises hippocampal function. A partial recovery of hippocampal synaptic plasticity emerges in advanced adulthood, however.
    DOI:  https://doi.org/10.1038/s41598-022-13127-y
  85. Genes Dev. 2022 Jun 02.
    The SpoVA membrane complex is required for dipicolinic acid import during sporulation and export during germination.
    Yongqiang Gao, Rocio Del Carmen Barajas-Ornelas, Jeremy D Amon, Fernando H Ramírez-Guadiana, Assaf Alon, Kelly P Brock, Debora S Marks, Andrew C Kruse, David Z Rudner.
      In response to starvation, endospore-forming bacteria differentiate into stress-resistant spores that can remain dormant for years yet rapidly germinate and resume growth in response to nutrients. The small molecule dipicolinic acid (DPA) plays a central role in both the stress resistance of the dormant spore and its exit from dormancy during germination. The spoVA locus is required for DPA import during sporulation and has been implicated in its export during germination, but the molecular bases are unclear. Here, we define the minimal set of proteins encoded in the Bacillus subtilis spoVA operon required for DPA import and demonstrate that these proteins form a membrane complex. Structural modeling of these components combined with mutagenesis and in vivo analysis reveal that the C and Eb subunits form a membrane channel, while the D subunit functions as a cytoplasmic plug. We show that point mutations that impair the interactions between D and the C-Eb membrane complex reduce the efficiency of DPA import during sporulation and reciprocally accelerate DPA release during germination. Our data support a model in which DPA transport into spores involves cycles of unplugging and then replugging the C-Eb membrane channel, while nutrient detection during germination triggers DPA release by unplugging it.
    Keywords:  DPA; dipicolinic acid; dormancy; germination; spoVA; sporulation
    DOI:  https://doi.org/10.1101/gad.349488.122
  86. Cell Rep. 2022 May 31. pii: S2211-1247(22)00652-0. [Epub ahead of print]39(9): 110877
    Mapping cis-regulatory elements in human neurons links psychiatric disease heritability and activity-regulated transcriptional programs.
    Carlos Sanchez-Priego, Ruiqi Hu, Linda L Boshans, Matthew Lalli, Justyna A Janas, Sarah E Williams, Zhiqiang Dong, Nan Yang.
      Genome-wide association studies (GWASs) have identified hundreds of loci associated with psychiatric diseases, yet there is a lack of understanding of disease pathophysiology. Common risk variants can shed light on the underlying molecular mechanisms; however, identifying causal variants remains challenging. We map cis-regulatory elements in human neurons derived from pluripotent stem cells. This system allows us to determine enhancers that activate the transcription of neuronal activity-regulated gene programs, which are thought to be critical for synaptic plasticity and are not possible to identify from postmortem tissues. Using the activity-by-contact model, we create variant-to-gene maps to interpret the function of GWAS variants. Our work nominates a subset of variants to elucidate the molecular mechanisms involving GWAS-significant loci. It also highlights that in vitro human cellular models are a powerful platform for identifying and mechanistic studies of human trait-associated genetic variants in cell states that are inaccessible from other types of human samples.
    Keywords:  CP: Molecular biology; CP: Neuroscience; cis-regulatory elements; human stem cells; neurons; non-coding variation; psychiatric diseases
    DOI:  https://doi.org/10.1016/j.celrep.2022.110877
  87. Nature. 2022 06;606(7912): 9
    Fix the process that led to Alzheimer's drug fiasco.
    Jason Karlawish.
      
    Keywords:  Medical research; Policy
    DOI:  https://doi.org/10.1038/d41586-022-01507-3
  88. Nat Biotechnol. 2022 Jun 02.
    Spatiotemporal multiplexed immunofluorescence imaging of living cells and tissues with bioorthogonal cycling of fluorescent probes.
    Jina Ko, Martin Wilkovitsch, Juhyun Oh, Rainer H Kohler, Evangelia Bolli, Mikael J Pittet, Claudio Vinegoni, David B Sykes, Hannes Mikula, Ralph Weissleder, Jonathan C T Carlson.
      Cells in complex organisms undergo frequent functional changes, but few methods allow comprehensive longitudinal profiling of living cells. Here we introduce scission-accelerated fluorophore exchange (SAFE), a method for multiplexed temporospatial imaging of living cells with immunofluorescence. SAFE uses a rapid bioorthogonal click chemistry to remove immunofluorescent signals from the surface of labeled cells, cycling the nanomolar-concentration reagents in seconds and enabling multiple rounds of staining of the same samples. It is non-toxic and functional in both dispersed cells and intact living tissues. We demonstrate multiparameter (n ≥ 14), non-disruptive imaging of murine peripheral blood mononuclear and bone marrow cells to profile cellular differentiation. We also show longitudinal multiplexed imaging of bone marrow progenitor cells as they develop into neutrophils over 6 days and real-time multiplexed cycling of living mouse hepatic tissues. We anticipate that SAFE will find broad utility for investigating physiologic dynamics in living systems.
    DOI:  https://doi.org/10.1038/s41587-022-01339-6
  89. Am J Dermatopathol. 2022 May 30.
    Shared Clonal Origin of Multiple Histiocytic and Dendritic Neoplasms and Polycythemia Vera, Follicular Lymphoma in 1 Patient.
    Cecilia Larocca, Vignesh Shanmugam, David Zemmour, Joseph H Antin, Andrew A Lane.
      
    DOI:  https://doi.org/10.1097/DAD.0000000000002231
  90. Int J Biol Sci. 2022 ;18(8): 3178-3193
    Sublytic C5b-9 Induces CCL3/4 Production and Macrophage Accumulation in Thy-1N Rats via PKC-α/p65/IRF-8 Axis.
    Wenbo Wang, Baomei Qian, Chenhui Zhao, Mingyu Peng, Longfei Liu, Mengxiao Xie, Na Peng, Qingling He, Shuai Ying, Yufeng Zhu, Tao Wang, Dajun Hu, Dan Zhao, Jing Zhang, Yingwei Wang, Wen Qiu.
      Mesangioproliferative glomerulonephritis (MsPGN) is a common human kidney disease. Rat Thy-1 nephritis (Thy-1N) is an animal model widely used for the study of MsPGN. Thy-1N is not only sublytic C5b-9-dependent, but also related to pro-inflammatory cytokine production and macrophage (Mφ) accumulation in rat renal tissues. In this study, we found that the expression or phosphorylation of chemokine CCL3/4, CD68 (Mφ marker), IRF-8, PKC-α and NF-κB-p65 (p65) were all up-regulated both in the renal tissues of Thy-1N rats (in vivo) and in the glomerular mesangial cells (GMCs) upon sublytic C5b-9 stimulation (in vitro). Further experiments in vitro revealed that the phosphorylated PKC-α (p-PKC-α) could promote p65 phosphorylation, and then p-p65 enhanced IRF-8 expression through binding to IRF-8 promotor (-591 ~ -582 nt and -299 ~ -290 nt). Additionally, up-regulation or silencing of IRF-8 gene promoted or reduced CCL3/4 production, and then regulated Mφ chemotaxis. The underlying mechanism involved in IRF-8 binding to CCL3 promoter (-249 ~ -236 nt), which resulted in CCL3 gene transcription. The experiments in vivo showed that knockdown of renal PKC-α, p65, IRF-8 and CCL3/4 genes could inhibit CCL3/4 production, Mφ accumulation, GMC proliferation and proteinuria of Thy-1N rats. Furthermore, p-PKC-α, p-p65, IRF-8, CCL3/4 expression and Mφ accumulation were also increased in the renal tissues of MsPGN patients. Collectively, these findings indicate that sublytic C5b-9 induces CCL3/4 production and Mφ accumulation via PKC-α/p65/IRF-8 axis, and finally aggravates the pathological changes of MsPGN.
    Keywords:  CCL3/4; IRF-8; Thy-1 nephritis; macrophage; mesangioproliferative glomerulonephritis; sublytic C5b-9
    DOI:  https://doi.org/10.7150/ijbs.69652
  91. Cell Host Microbe. 2022 May 26. pii: S1931-3128(22)00263-3. [Epub ahead of print]
    Impact of a 7-day homogeneous diet on interpersonal variation in human gut microbiomes and metabolomes.
    Leah Guthrie, Sean Paul Spencer, Dalia Perelman, Will Van Treuren, Shuo Han, Feiqiao Brian Yu, Erica D Sonnenburg, Michael A Fischbach, Timothy W Meyer, Justin L Sonnenburg.
      Gut microbiota metabolism of dietary compounds generates a vast array of microbiome-dependent metabolites (MDMs), which are highly variable between individuals. The uremic MDMs (uMDMs) phenylacetylglutamine (PAG), p-cresol sulfate (PCS), and indoxyl sulfate (IS) accumulate during renal failure and are associated with poor outcomes. Targeted dietary interventions may reduce toxic MDM generation; however, it is unclear if inter-individual differences in diet or gut microbiome dominantly contribute to MDM variance. Here, we use a 7-day homogeneous average American diet to standardize dietary precursor availability in 21 healthy individuals. During dietary homogeneity, the coefficient of variation in PAG, PCS, and IS (primary outcome) did not decrease, nor did inter-individual variation in most identified metabolites; other microbiome metrics showed no or modest responses to the intervention. Host identity and age are dominant contributors to variability in MDMs. These results highlight the potential need to pair dietary modification with microbial therapies to control MDM profiles.
    Keywords:  dietary intervention; microbiome; microbiome-dependent metabolites; precision medicine; uremic solutes
    DOI:  https://doi.org/10.1016/j.chom.2022.05.003
  92. Nat Commun. 2022 May 31. 13(1): 3012
    Structural variant-based pangenome construction has low sensitivity to variability of haplotype-resolved bovine assemblies.
    Alexander S Leonard, Danang Crysnanto, Zih-Hua Fang, Michael P Heaton, Brian L Vander Ley, Carolina Herrera, Heinrich Bollwein, Derek M Bickhart, Kristen L Kuhn, Timothy P L Smith, Benjamin D Rosen, Hubert Pausch.
      Advantages of pangenomes over linear reference assemblies for genome research have recently been established. However, potential effects of sequence platform and assembly approach, or of combining assemblies created by different approaches, on pangenome construction have not been investigated. Here we generate haplotype-resolved assemblies from the offspring of three bovine trios representing increasing levels of heterozygosity that each demonstrate a substantial improvement in contiguity, completeness, and accuracy over the current Bos taurus reference genome. Diploid coverage as low as 20x for HiFi or 60x for ONT is sufficient to produce two haplotype-resolved assemblies meeting standards set by the Vertebrate Genomes Project. Structural variant-based pangenomes created from the haplotype-resolved assemblies demonstrate significant consensus regardless of sequence platform, assembler algorithm, or coverage. Inspecting pangenome topologies identifies 90 thousand structural variants including 931 overlapping with coding sequences; this approach reveals variants affecting QRICH2, PRDM9, HSPA1A, TAS2R46, and GC that have potential to affect phenotype.
    DOI:  https://doi.org/10.1038/s41467-022-30680-2
  93. Pediatrics. 2022 Jun 01. pii: e2021054952. [Epub ahead of print]149(6):
    Long-Term Care for Children With Medical Complexity: A Call for Data.
    Leigh Alon, Renee D Boss, Rebecca Seltzer.
      
    DOI:  https://doi.org/10.1542/peds.2021-054952
  94. Med (N Y). 2022 May 26. pii: S2666-6340(22)00184-2. [Epub ahead of print]
    Cross-tissue, single-cell stromal atlas identifies shared pathological fibroblast phenotypes in four chronic inflammatory diseases.
    Ilya Korsunsky, Kevin Wei, Mathilde Pohin, Edy Y Kim, Francesca Barone, Triin Major, Emily Taylor, Rahul Ravindran, Samuel Kemble, Gerald F M Watts, A Helena Jonsson, Yunju Jeong, Humra Athar, Dylan Windell, Joyce B Kang, Matthias Friedrich, Jason Turner, Saba Nayar, Benjamin A Fisher, Karim Raza, Jennifer L Marshall, Adam P Croft, Tomoyoshi Tamura, Lynette M Sholl, Marina Vivero, Ivan O Rosas, Simon J Bowman, Mark Coles, Andreas P Frei, Kara Lassen, Andrew Filer, Fiona Powrie, Christopher D Buckley, Michael B Brenner, Soumya Raychaudhuri.
       BACKGROUND: Pro-inflammatory fibroblasts are critical for pathogenesis in rheumatoid arthritis, inflammatory bowel disease, interstitial lung disease, and Sjögren's syndrome and represent a novel therapeutic target for chronic inflammatory disease. However, the heterogeneity of fibroblast phenotypes, exacerbated by the lack of a common cross-tissue taxonomy, has limited our understanding of which pathways are shared by multiple diseases.
    METHODS: We profiled fibroblasts derived from inflamed and non-inflamed synovium, intestine, lungs, and salivary glands from affected individuals with single-cell RNA sequencing. We integrated all fibroblasts into a multi-tissue atlas to characterize shared and tissue-specific phenotypes.
    FINDINGS: Two shared clusters, CXCL10+CCL19+ immune-interacting and SPARC+COL3A1+ vascular-interacting fibroblasts, were expanded in all inflamed tissues and mapped to dermal analogs in a public atopic dermatitis atlas. We confirmed these human pro-inflammatory fibroblasts in animal models of lung, joint, and intestinal inflammation.
    CONCLUSIONS: This work represents a thorough investigation into fibroblasts across organ systems, individual donors, and disease states that reveals shared pathogenic activation states across four chronic inflammatory diseases.
    FUNDING: Grant from F. Hoffmann-La Roche (Roche) AG.
    Keywords:  Foundational research; Sjögren's syndrome; atlas; fibroblasts; inflammation; integration; interstitial lung disease; rheumatoid arthritis; scRNA-seq; stromal; ulcerative colitis
    DOI:  https://doi.org/10.1016/j.medj.2022.05.002
  95. Nat Commun. 2022 May 30. 13(1): 3010
    Myriapod genomes reveal ancestral horizontal gene transfer and hormonal gene loss in millipedes.
    Wai Lok So, Wenyan Nong, Yichun Xie, Tobias Baril, Hai-Yao Ma, Zhe Qu, Jasmine Haimovitz, Thomas Swale, Juan Diego Gaitan-Espitia, Kwok Fai Lau, Stephen S Tobe, William G Bendena, Zhen-Peng Kai, Alexander Hayward, Jerome H L Hui.
      Animals display a fascinating diversity of body plans. Correspondingly, genomic analyses have revealed dynamic evolution of gene gains and losses among animal lineages. Here we sequence six new myriapod genomes (three millipedes, three centipedes) at key phylogenetic positions within this major but understudied arthropod lineage. We combine these with existing genomic resources to conduct a comparative analysis across all available myriapod genomes. We find that millipedes generally have considerably smaller genomes than centipedes, with the repeatome being a major contributor to genome size, driven by independent large gains of transposons in three centipede species. In contrast to millipedes, centipedes gained a large number of gene families after the subphyla diverged, with gains contributing to sensory and locomotory adaptations that facilitated their ecological shift to predation. We identify distinct horizontal gene transfer (HGT) events from bacteria to millipedes and centipedes, with no identifiable HGTs shared among all myriapods. Loss of juvenile hormone O-methyltransferase, a key enzyme in catalysing sesquiterpenoid hormone production in arthropods, was also revealed in all millipede lineages. Our findings suggest that the rapid evolution of distinct genomic pathways in centipede and millipede lineages following their divergence from the myriapod ancestor, was shaped by differing ecological pressures.
    DOI:  https://doi.org/10.1038/s41467-022-30690-0
  96. Nat Commun. 2022 Jun 01. 13(1): 3056
    Nuclear Vav3 is required for polycomb repression complex-1 activity in B-cell lymphoblastic leukemogenesis.
    R C Nayak, K H Chang, A K Singh, M Kotliar, M Desai, A M Wellendorf, M Wunderlich, J Bartram, B Mizukawa, M Cuadrado, P Dexheimer, A Barski, X R Bustelo, N N Nassar, J A Cancelas.
      Acute B-cell lymphoblastic leukemia (B-ALL) results from oligo-clonal evolution of B-cell progenitors endowed with initiating and propagating leukemia properties. The activation of both the Rac guanine nucleotide exchange factor (Rac GEF) Vav3 and Rac GTPases is required for leukemogenesis mediated by the oncogenic fusion protein BCR-ABL. Vav3 expression becomes predominantly nuclear upon expression of BCR-ABL signature. In the nucleus, Vav3 interacts with BCR-ABL, Rac, and the polycomb repression complex (PRC) proteins Bmi1, Ring1b and Ezh2. The GEF activity of Vav3 is required for the proliferation, Bmi1-dependent B-cell progenitor self-renewal, nuclear Rac activation, protein interaction with Bmi1, mono-ubiquitination of H2A(K119) (H2AK119Ub) and repression of PRC-1 (PRC1) downstream target loci, of leukemic B-cell progenitors. Vav3 deficiency results in de-repression of negative regulators of cell proliferation and repression of oncogenic transcriptional factors. Mechanistically, we show that Vav3 prevents the Phlpp2-sensitive and Akt (S473)-dependent phosphorylation of Bmi1 on the regulatory residue S314 that, in turn, promotes the transcriptional factor reprogramming of leukemic B-cell progenitors. These results highlight the importance of non-canonical nuclear Rho GTPase signaling in leukemogenesis.
    DOI:  https://doi.org/10.1038/s41467-022-30651-7
  97. Proc Natl Acad Sci U S A. 2022 Jun 07. 119(23): e2203965119
    The nonclassical MHC class I Qa-1 expressed in layer 6 neurons regulates activity-dependent plasticity via microglial CD94/NKG2 in the cortex.
    Ioana A Marin, Alan Y Gutman-Wei, Kylie S Chew, Aram J Raissi, Maja Djurisic, Carla J Shatz.
      Significance Molecules regulated by neuronal activity are necessary for circuits to adapt to changing inputs. Specific classical major histocompatibility class I (MHCI) molecules play roles in circuit and synaptic plasticity, but the function of most members of this family remains unexplored in brain. Here, we show that a nonclassical MHCI molecule, Qa-1 (H2-T23), is expressed in a subset of excitatory neurons and regulated by visually driven activity in the cerebral cortex. Moreover, CD94/NKG2 heterodimers, cognate receptors for Qa-1, are expressed in microglia. A functional interaction between Qa-1 and CD94/NKG2 is necessary for regulating the magnitude of ocular dominance plasticity during the critical period in the visual cortex, implying an interaction in which activity-dependent changes in neurons may be monitored by microglia.
    Keywords:  corticothalamic neurons; microglial innate immune receptor; neuronal MHCI; ocular dominance plasticity
    DOI:  https://doi.org/10.1073/pnas.2203965119
  98. Nat Commun. 2022 Jun 01. 13(1): 3041
    Cryo-EM structures of Gid12-bound GID E3 reveal steric blockade as a mechanism inhibiting substrate ubiquitylation.
    Shuai Qiao, Chia-Wei Lee, Dawafuti Sherpa, Jakub Chrustowicz, Jingdong Cheng, Maximilian Duennebacke, Barbara Steigenberger, Ozge Karayel, Duc Tung Vu, Susanne von Gronau, Matthias Mann, Florian Wilfling, Brenda A Schulman.
      Protein degradation, a major eukaryotic response to cellular signals, is subject to numerous layers of regulation. In yeast, the evolutionarily conserved GID E3 ligase mediates glucose-induced degradation of fructose-1,6-bisphosphatase (Fbp1), malate dehydrogenase (Mdh2), and other gluconeogenic enzymes. "GID" is a collection of E3 ligase complexes; a core scaffold, RING-type catalytic core, and a supramolecular assembly module together with interchangeable substrate receptors select targets for ubiquitylation. However, knowledge of additional cellular factors directly regulating GID-type E3s remains rudimentary. Here, we structurally and biochemically characterize Gid12 as a modulator of the GID E3 ligase complex. Our collection of cryo-EM reconstructions shows that Gid12 forms an extensive interface sealing the substrate receptor Gid4 onto the scaffold, and remodeling the degron binding site. Gid12 also sterically blocks a recruited Fbp1 or Mdh2 from the ubiquitylation active sites. Our analysis of the role of Gid12 establishes principles that may more generally underlie E3 ligase regulation.
    DOI:  https://doi.org/10.1038/s41467-022-30803-9
  99. J Clin Invest. 2022 Jun 01. pii: e161661. [Epub ahead of print]132(11):
    A conversation with Elizabeth Blackburn and Viviane Tabar.
    Inés Fernández Maestre, Yanyang Chen, Brianna Naizir, Ushma S Neill.
      
    DOI:  https://doi.org/10.1172/JCI161661
  100. Front Immunol. 2022 ;13 880829
    Neutralizing Antibodies Against Factor VIII Can Occur Through a Non-Germinal Center Pathway.
    Seema R Patel, Taran S Lundgren, Wallace Hunter Baldwin, Courtney Cox, Ernest T Parker, John F Healey, Ryan P Jajosky, Patricia E Zerra, Cassandra D Josephson, Christopher B Doering, Sean R Stowell, Shannon L Meeks.
      Humoral immunity to factor VIII (FVIII) represents a significant challenge for the treatment of patients with hemophilia A. Current paradigms indicate that neutralizing antibodies against FVIII (inhibitors) occur through a classical CD4 T cell, germinal center (GC) dependent process. However, clinical observations suggest that the nature of the immune response to FVIII may differ between patients. While some patients produce persistent low or high inhibitor titers, others generate a transient response. Moreover, FVIII reactive memory B cells are only detectable in some patients with sustained inhibitor titers. The determinants regulating the type of immune response a patient develops, let alone how the immune response differs in these patients remains incompletely understood. One hypothesis is that polymorphisms within immunoregulatory genes alter the underlying immune response to FVIII, and thereby the inhibitor response. Consistent with this, studies report that inhibitor titers to FVIII differ in animals with the same F8 pathogenic variant but completely distinct backgrounds; though, how these genetic disparities affect the immune response to FVIII remains to be investigated. Given this, we sought to mechanistically dissect how genetics impact the underlying immune response to FVIII. In particular, as the risk of producing inhibitors is weakly associated with differences in HLA, we hypothesized that genetic factors other than HLA influence the immune response to FVIII and downstream inhibitor formation. Our data demonstrate that FVIII deficient mice encoding the same MHC and F8 variant produce disparate inhibitor titers, and that the type of inhibitor response formed associates with the ability to generate GCs. Interestingly, the formation of antibodies through a GC or non-GC pathway does not appear to be due to differences in CD4 T cell immunity, as the CD4 T cell response to an immunodominant epitope in FVIII was similar in these mice. These results indicate that genetics can impact the process by which inhibitors develop and may in part explain the apparent propensity of patients to form distinct inhibitor responses. Moreover, these data highlight an underappreciated immunological pathway of humoral immunity to FVIII and lay the groundwork for identification of biomarkers for the development of approaches to tolerize against FVIII.
    Keywords:  B cells; extrafollicular pathway; germinal center; hemophilia A; neutralizing antibodies (inhibitors)
    DOI:  https://doi.org/10.3389/fimmu.2022.880829
  101. Mol Biol Cell. 2022 Jun 02. mbcE22020032
    Cardiac inducing colonies halt fibroblast activation and induce cardiac/endothelial cells to move and expand via paracrine signaling.
    Samiksha Mahapatra, Michael V R Sharma, Breanna Brownson, Vaughn E Gallicano, G Ian Gallicano.
      Myocardial fibrosis (MF), a common event that develops after myocardial infarction, initially is a reparative process, but eventually leads to heart failure (HF) and sudden cardiac arrest (SCA). In MF, the infarct area is replaced by a collagenous-based scar induced by 'excessive' collagen deposition from activated cardiac fibroblasts. The scar prevents ventricular wall thinning; however, over time it expands to non-infarcted myocardium. Therapies to prevent fibrosis include reperfusion, anti-fibrotic agents, and ACE inhibitors. Paracrine factor (PF)/stem cell research has recently gained significance as a therapy. We consistently find that cardiac inducing colonies (CiCs) [derived from human germline pluripotent stem cell (hgPSC)] secrete PFs at physiologically relevant concentrations that suppress cardiac fibroblast (HCF) activation and excessive ECM protein secretion. These factors also affect human cardiomyocytes and endothelial cells by inducing migration/proliferation of both populations into a myocardial wound model. Finally, CiC factors modulate matrix turnover and pro-inflammation. Taken together, we show that CiCs could help tip the balance from fibrosis towards repair.
    DOI:  https://doi.org/10.1091/mbc.E22-02-0032
  102. Nat Commun. 2022 May 31. 13(1): 3018
    Blocking phospholamban with VHH intrabodies enhances contractility and relaxation in heart failure.
    Erwin De Genst, Kylie S Foo, Yao Xiao, Eduarde Rohner, Emma de Vries, Jesper Sohlmér, Nevin Witman, Alejandro Hidalgo, Terje R S Kolstad, William E Louch, Susanne Pehrsson, Andrew Park, Yasuhiro Ikeda, Xidan Li, Lorenz M Mayr, Kate Wickson, Karin Jennbacken, Kenny Hansson, Regina Fritsche-Danielson, James Hunt, Kenneth R Chien.
      The dysregulated physical interaction between two intracellular membrane proteins, the sarco/endoplasmic reticulum Ca2+ ATPase and its reversible inhibitor phospholamban, induces heart failure by inhibiting calcium cycling. While phospholamban is a bona-fide therapeutic target, approaches to selectively inhibit this protein remain elusive. Here, we report the in vivo application of intracellular acting antibodies (intrabodies), derived from the variable domain of camelid heavy-chain antibodies, to modulate the function of phospholamban. Using a synthetic VHH phage-display library, we identify intrabodies with high affinity and specificity for different conformational states of phospholamban. Rapid phenotypic screening, via modified mRNA transfection of primary cells and tissue, efficiently identifies the intrabody with most desirable features. Adeno-associated virus mediated delivery of this intrabody results in improvement of cardiac performance in a murine heart failure model. Our strategy for generating intrabodies to investigate cardiac disease combined with modified mRNA and adeno-associated virus screening could reveal unique future therapeutic opportunities.
    DOI:  https://doi.org/10.1038/s41467-022-29703-9
  103. Nat Methods. 2022 Jun 02.
    Identification of cell types in multiplexed in situ images by combining protein expression and spatial information using CELESTA.
    Weiruo Zhang, Irene Li, Nathan E Reticker-Flynn, Zinaida Good, Serena Chang, Nikolay Samusik, Saumyaa Saumyaa, Yuanyuan Li, Xin Zhou, Rachel Liang, Christina S Kong, Quynh-Thu Le, Andrew J Gentles, John B Sunwoo, Garry P Nolan, Edgar G Engleman, Sylvia K Plevritis.
      Advances in multiplexed in situ imaging are revealing important insights in spatial biology. However, cell type identification remains a major challenge in imaging analysis, with most existing methods involving substantial manual assessment and subjective decisions for thousands of cells. We developed an unsupervised machine learning algorithm, CELESTA, which identifies the cell type of each cell, individually, using the cell's marker expression profile and, when needed, its spatial information. We demonstrate the performance of CELESTA on multiplexed immunofluorescence images of colorectal cancer and head and neck squamous cell carcinoma (HNSCC). Using the cell types identified by CELESTA, we identify tissue architecture associated with lymph node metastasis in HNSCC, and validate our findings in an independent cohort. By coupling our spatial analysis with single-cell RNA-sequencing data on proximal sections of the same specimens, we identify cell-cell crosstalk associated with lymph node metastasis, demonstrating the power of CELESTA to facilitate identification of clinically relevant interactions.
    DOI:  https://doi.org/10.1038/s41592-022-01498-z
  104. Nat Rev Immunol. 2022 Jun 01.
    Role of thymic stromal lymphopoietin in allergy and beyond.
    Risa Ebina-Shibuya, Warren J Leonard.
      Thymic stromal lymphopoietin (TSLP) is a pleiotropic cytokine that acts on multiple cell lineages, including dendritic cells, T cells, B cells, neutrophils, mast cells, eosinophils and innate lymphoid cells, affecting their maturation, survival and recruitment. It is best known for its role in promoting type 2 immune responses such as in allergic diseases and, in 2021, a monoclonal antibody targeting TSLP was approved for the treatment of severe asthma. However, it is now clear that TSLP has many other important roles in a variety of settings. Indeed, several genetic variants for TSLP are linked to disease severity, and chromosomal alterations in TSLP are common in certain cancers, indicating important roles of TSLP in disease. In this Review, we discuss recent advances in TSLP biology, highlighting how it regulates the tissue environment not only in allergic disease but also in infectious diseases, inflammatory diseases and cancer. Encouragingly, therapies targeting the TSLP pathway are being actively pursued for several diseases.
    DOI:  https://doi.org/10.1038/s41577-022-00735-y
  105. mBio. 2022 May 31. e0127022
    The Shigella Spp. Type III Effector Protein OspB Is a Cysteine Protease.
    Thomas E Wood, Kathleen A Westervelt, Jessica M Yoon, Heather D Eshleman, Roie Levy, Henry Burnes, Daniel J Slade, Cammie F Lesser, Marcia B Goldberg.
      The type III secretion system is required for virulence of many pathogenic bacteria. Bacterial effector proteins delivered into target host cells by this system modulate host signaling pathways and processes in a manner that promotes infection. Here, we define the activity of the effector protein OspB of the human pathogen Shigella spp., the etiological agent of shigellosis and bacillary dysentery. Using the yeast Saccharomyces cerevisiae as a model organism, we show that OspB sensitizes cells to inhibition of TORC1, the central regulator of growth and metabolism. In silico analyses reveal that OspB bears structural homology to bacterial cysteine proteases that target mammalian cell processes, and we define a conserved cysteine-histidine catalytic dyad required for OspB function. Using yeast genetic screens, we identify a crucial role for the arginine N-degron pathway in the yeast growth inhibition phenotype and show that inositol hexakisphosphate is an OspB cofactor. We find that a yeast substrate for OspB is the TORC1 component Tco89p, proteolytic cleavage of which generates a C-terminal fragment that is targeted for degradation via the arginine N-degron pathway; processing and degradation of Tco89p is required for the OspB phenotype. In all, we demonstrate that the Shigella T3SS effector OspB is a cysteine protease and decipher its interplay with eukaryotic cell processes. IMPORTANCE Shigella spp. are important human pathogens and among the leading causes of diarrheal mortality worldwide, especially in children. Virulence depends on the Shigella type III secretion system (T3SS). Definition of the roles of the bacterial effector proteins secreted by the T3SS is key to understanding Shigella pathogenesis. The effector protein OspB contributes to a range of phenotypes during infection, yet the mechanism of action is unknown. Here, we show that S. flexneri OspB possesses cysteine protease activity in both yeast and mammalian cells, and that enzymatic activity of OspB depends on a conserved cysteine-histidine catalytic dyad. We determine how its protease activity sensitizes cells to TORC1 inhibition in yeast, finding that OspB cleaves a component of yeast TORC1, and that the degradation of the C-terminal cleavage product is responsible for OspB-mediated hypersensitivity to TORC1 inhibitors. Thus, OspB is a cysteine protease that depends on a conserved cysteine-histidine catalytic dyad.
    Keywords:  OspB; Shigella; TORC1; Tco89p; catalytic dyad; cysteine protease; inositol hexakisphosphate
    DOI:  https://doi.org/10.1128/mbio.01270-22
  106. Nat Genet. 2022 Jun 02.
    Genetic correlates of phenotypic heterogeneity in autism.
    EU-AIMS LEAP
      The substantial phenotypic heterogeneity in autism limits our understanding of its genetic etiology. To address this gap, here we investigated genetic differences between autistic individuals (nmax = 12,893) based on core and associated features of autism, co-occurring developmental disabilities and sex. We conducted a comprehensive factor analysis of core autism features in autistic individuals and identified six factors. Common genetic variants were associated with the core factors, but de novo variants were not. We found that higher autism polygenic scores (PGS) were associated with lower likelihood of co-occurring developmental disabilities in autistic individuals. Furthermore, in autistic individuals without co-occurring intellectual disability (ID), autism PGS are overinherited by autistic females compared to males. Finally, we observed higher SNP heritability for autistic males and for autistic individuals without ID. Deeper phenotypic characterization will be critical in determining how the complex underlying genetics shape cognition, behavior and co-occurring conditions in autism.
    DOI:  https://doi.org/10.1038/s41588-022-01072-5
  107. Cell Rep. 2022 May 31. pii: S2211-1247(22)00677-5. [Epub ahead of print]39(9): 110902
    An integrated model for Gpr124 function in Wnt7a/b signaling among vertebrates.
    Michelle America, Naguissa Bostaille, Marie Eubelen, Maud Martin, Didier Y R Stainier, Benoit Vanhollebeke.
      Within the central nervous system, Wnt7a/b are unambiguously discriminated from other Wnt ligands by an endothelial receptor complex made of the glycosylphosphatidylinositol (GPI)-anchored Reck and the adhesion G protein-coupled receptor (GPCR) Gpr124. Reck is a Wnt7a/b-specific receptor, while Gpr124 facilitates the delivery of Reck-bound Wnt7a/b ligands to Frizzled, through partially characterized mechanisms. We report that, in zebrafish, the Gpr124-Frizzled interactions are dominated by intracellular scaffolds that exploit the striking molecular mimicry between Gpr124 and Frizzled intracellular domains (ICDs): an internal Dvl-binding motif and a C-terminal ETTV motif that recruits Dlg4 and Magi3. By contrast, mammalian Gpr124 receptors exhibit an ICD-independent interaction mechanism governed by species-specific attributes of their transmembrane and extracellular domains. This mechanism seemingly evolved to replace the Dvl-mediated mechanism. By contrasting zebrafish, mouse, and human Gpr124, this study provides insights into the evolution of Gpr124/Reck function across the vertebrate clade, a receptor complex uniquely implicated in Wnt ligand-specific cellular responses.
    Keywords:  BBB; CP: Cell biology; Dlg4; Gpr124; Magi3; Reck; Wnt/β-catenin signaling; Wnt7a; Wnt7b; blood-brain barrier; brain angiogenesis
    DOI:  https://doi.org/10.1016/j.celrep.2022.110902
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