bims-nimamd Biomed News
on Neuroimmunity and neuroinflammation in ageing and metabolic disease
Issue of 2023–02–12
sixty-four papers selected by
Fawaz Alzaïd, Sorbonne Université
  1. ESCRT-dependent STING degradation inhibits steady-state and cGAMP-induced signalling.
  2. The p97-UBXD8 complex regulates ER-Mitochondria contact sites by altering membrane lipid saturation and composition.
  3. Widespread contribution of transposable elements to the rewiring of mammalian 3D genomes.
  4. Temporal-iCLIP captures co-transcriptional RNA-protein interactions.
  5. An updated suite of viral vectors for in vivo calcium imaging using intracerebral and retro-orbital injections in male mice.
  6. Myristic acid as a checkpoint to regulate STING-dependent autophagy and interferon responses by promoting N-myristoylation.
  7. TRIM40 is a pathogenic driver of inflammatory bowel disease subverting intestinal barrier integrity.
  8. Splicing factor SRSF1 deficiency in the liver triggers NASH-like pathology and cell death.
  9. Human IL-23 is essential for IFN-γ-dependent immunity to mycobacteria.
  10. H2B ubiquitination recruits FACT to maintain a stable altered nucleosome state for transcriptional activation.
  11. LSD1/PRMT6-targeting gene therapy to attenuate androgen receptor toxic gain-of-function ameliorates spinobulbar muscular atrophy phenotypes in flies and mice.
  12. Adipocyte-derived extracellular vesicles increase insulin secretion through transport of insulinotropic protein cargo.
  13. Hepatocytes demarcated by EphB2 contribute to the progression of nonalcoholic steatohepatitis.
  14. Lipid-droplet associated mitochondria promote fatty-acid oxidation through a distinct bioenergetic pattern in male Wistar rats.
  15. Extending genetic risk for Alzheimer's disease from host to holobiont.
  16. ZBTB12 is a molecular barrier to dedifferentiation in human pluripotent stem cells.
  17. What no one told me.
  18. Application of high-throughput single-nucleus DNA sequencing in pancreatic cancer.
  19. DRAK2 contributes to type 1 diabetes by negatively regulating IL-2 sensitivity to alter regulatory T cell development.
  20. Recycling of modified H2A-H2B provides short-term memory of chromatin states.
  21. A conformational switch in clathrin light chain regulates lattice structure and endocytosis at the plasma membrane of mammalian cells.
  22. Oxidized mitochondrial DNA: a protective signal gone awry.
  23. Defective excitation-contraction coupling and mitochondrial respiration precede mitochondrial Ca2+ accumulation in spinobulbar muscular atrophy skeletal muscle.
  24. Light-driven biological actuators to probe the rheology of 3D microtissues.
  25. Histone variant H2A.Z modulates nucleosome dynamics to promote DNA accessibility.
  26. Microbiota-derived acetate enhances host antiviral response via NLRP3.
  27. Cryo-EM structure of the RADAR supramolecular anti-phage defense complex.
  28. Cartography of Genomic Interactions Enables Deep Analysis of Single-Cell Expression Data.
  29. The endogenous repertoire harbors self-reactive CD4+ T cell clones that adopt a follicular helper T cell-like phenotype at steady state.
  30. Melatonin protects against cadmium-induced oxidative stress via mitochondrial STAT3 signaling in human prostate stromal cells.
  31. Peanut allergen inhibition prevents anaphylaxis in a humanized mouse model.
  32. A cis-regulatory lexicon of DNA motif combinations mediating cell-type-specific gene regulation.
  33. Hippocampal-cortical coupling differentiates long-term memory processes.
  34. Limited Proteolysis-Mass Spectrometry Reveals Aging-Associated Changes in Cerebrospinal Fluid Protein Abundances and Structures.
  35. TET2 guards against unchecked BATF3-induced CAR T cell expansion.
  36. Mitochondrial fragmentation and donut formation enhance mitochondrial secretion to promote osteogenesis.
  37. Cell surface-bound La protein regulates the cell fusion stage of osteoclastogenesis.
  38. CryoET reveals organelle phenotypes in huntington disease patient iPSC-derived and mouse primary neurons.
  39. Metabolic reprogramming by immune-responsive gene 1 up-regulation improves donor heart preservation and function.
  40. Polygenic Scores Help Reduce Racial Disparities in Predictive Accuracy of Automated Type 1 Diabetes Classification Algorithms.
  41. Diverse CMT2 neuropathies are linked to aberrant G3BP interactions in stress granules.
  42. Different pathways for engulfment and endocytosis of liquid droplets by nanovesicles.
  43. Remodeling of colon plasma cell repertoire within ulcerative colitis patients.
  44. Structures of LRP2 reveal a molecular machine for endocytosis.
  45. Sodium perturbs mitochondrial respiration and induces dysfunctional Tregs.
  46. Exaggerated responses to a virus long gone.
  47. Actin cytoskeleton vulnerability to disulfide stress mediates disulfidptosis.
  48. Cpeb1b-mediated cytoplasmic polyadenylation of shha mRNA modulates zebrafish definitive hematopoiesis.
  49. Mechanisms of myeloid cell entry to the healthy and diseased central nervous system.
  50. An engineered variant of MECR reductase reveals indispensability of long-chain acyl-ACPs for mitochondrial respiration.
  51. An adipocentric perspective on the development and progression of non-alcoholic fatty liver disease.
  52. Structures of BIRC6-client complexes provide a mechanism of Smac-mediated release of caspases.
  53. How a tiny genetic change inflicts old age on young kids.
  54. Intrafusal-fiber LRP4 for muscle spindle formation and maintenance in adult and aged animals.
  55. Hedgehog is relayed through dynamic heparan sulfate interactions to shape its gradient.
  56. Bacteriophages inhibit and evade cGAS-like immune function in bacteria.
  57. Angiotensin-converting enzyme inhibitor promotes angiogenesis through Sp1/Sp3-mediated inhibition of notch signaling in male mice.
  58. Integrated cardio-behavioral responses to threat define defensive states.
  59. Salty Treg cells get out of balance.
  60. The cellular coding of temperature in the mammalian cortex.
  61. A CpG island-encoded mechanism protects genes from premature transcription termination.
  62. Human atlastins are sufficient to drive the fusion of liposomes with a physiological lipid composition.
  63. How fingerprints get their one-of-a-kind swirls.
  64. Smart microscopes spot fleeting biology.
  1. Nat Commun. 2023 Feb 04. 14(1): 611
    ESCRT-dependent STING degradation inhibits steady-state and cGAMP-induced signalling.
    Matteo Gentili, Bingxu Liu, Malvina Papanastasiou, Deborah Dele-Oni, Marc A Schwartz, Rebecca J Carlson, Aziz M Al'Khafaji, Karsten Krug, Adam Brown, John G Doench, Steven A Carr, Nir Hacohen.
      Stimulator of interferon genes (STING) is an intracellular sensor of cyclic di-nucleotides involved in the innate immune response against pathogen- or self-derived DNA. STING trafficking is tightly linked to its function, and its dysregulation can lead to disease. Here, we systematically characterize genes regulating STING trafficking and examine their impact on STING-mediated responses. Using proximity-ligation proteomics and genetic screens, we demonstrate that an endosomal sorting complex required for transport (ESCRT) complex containing HGS, VPS37A and UBAP1 promotes STING degradation, thereby terminating STING-mediated signaling. Mechanistically, STING oligomerization increases its ubiquitination by UBE2N, forming a platform for ESCRT recruitment at the endosome that terminates STING signaling via sorting in the lysosome. Finally, we show that expression of a UBAP1 mutant identified in patients with hereditary spastic paraplegia and associated with disrupted ESCRT function, increases steady-state STING-dependent type I IFN responses in healthy primary monocyte-derived dendritic cells and fibroblasts. Based on these findings, we propose that STING is subject to a tonic degradative flux and that the ESCRT complex acts as a homeostatic regulator of STING signaling.
    DOI:  https://doi.org/10.1038/s41467-023-36132-9
  2. Nat Commun. 2023 Feb 06. 14(1): 638
    The p97-UBXD8 complex regulates ER-Mitochondria contact sites by altering membrane lipid saturation and composition.
    Rakesh Ganji, Joao A Paulo, Yuecheng Xi, Ian Kline, Jiang Zhu, Christoph S Clemen, Conrad C Weihl, John G Purdy, Steve P Gygi, Malavika Raman.
      The intimate association between the endoplasmic reticulum (ER) and mitochondrial membranes at ER-Mitochondria contact sites (ERMCS) is a platform for critical cellular processes, particularly lipid synthesis. How contacts are remodeled and the impact of altered contacts on lipid metabolism remains poorly understood. We show that the p97 AAA-ATPase and its adaptor ubiquitin-X domain adaptor 8 (UBXD8) regulate ERMCS. The p97-UBXD8 complex localizes to contacts and its loss increases contacts in a manner that is dependent on p97 catalytic activity. Quantitative proteomics and lipidomics of ERMCS demonstrates alterations in proteins regulating lipid metabolism and a significant change in membrane lipid saturation upon UBXD8 deletion. Loss of p97-UBXD8 increased membrane lipid saturation via SREBP1 and the lipid desaturase SCD1. Aberrant contacts can be rescued by unsaturated fatty acids or overexpression of SCD1. We find that the SREBP1-SCD1 pathway is negatively impacted in the brains of mice with p97 mutations that cause neurodegeneration. We propose that contacts are exquisitely sensitive to alterations to membrane lipid composition and saturation.
    DOI:  https://doi.org/10.1038/s41467-023-36298-2
  3. Nat Commun. 2023 Feb 06. 14(1): 634
    Widespread contribution of transposable elements to the rewiring of mammalian 3D genomes.
    Mayank N K Choudhary, Kara Quaid, Xiaoyun Xing, Heather Schmidt, Ting Wang.
      Transposable elements (TEs) are major contributors of genetic material in mammalian genomes. These often include binding sites for architectural proteins, including the multifarious master protein, CTCF, which shapes the 3D genome by creating loops, domains, compartment borders, and RNA-DNA interactions. These play a role in the compact packaging of DNA and have the potential to facilitate regulatory function. In this study, we explore the widespread contribution of TEs to mammalian 3D genomes by quantifying the extent to which they give rise to loops and domain border differences across various cell types and species using several 3D genome mapping technologies. We show that specific families and subfamilies of TEs have contributed to lineage-specific 3D chromatin structures across mammalian species. In many cases, these loops may facilitate sustained interaction between distant cis-regulatory elements and target genes, and domains may segregate chromatin state to impact gene expression in a lineage-specific manner. An experimental validation of our analytical findings using CRISPR-Cas9 to delete a candidate TE resulted in disruption of species-specific 3D chromatin structure. Taken together, we comprehensively quantify and selectively validate our finding that TEs contribute to shaping 3D genome organization and may, in some cases, impact gene regulation during the course of mammalian evolution.
    DOI:  https://doi.org/10.1038/s41467-023-36364-9
  4. Nat Commun. 2023 Feb 08. 14(1): 696
    Temporal-iCLIP captures co-transcriptional RNA-protein interactions.
    Ross A Cordiner, Yuhui Dou, Rune Thomsen, Andrii Bugai, Sander Granneman, Torben Heick Jensen.
      Dynamic RNA-protein interactions govern the co-transcriptional packaging of RNA polymerase II (RNAPII)-derived transcripts. Yet, our current understanding of this process in vivo primarily stems from steady state analysis. To remedy this, we here conduct temporal-iCLIP (tiCLIP), combining RNAPII transcriptional synchronisation with UV cross-linking of RNA-protein complexes at serial timepoints. We apply tiCLIP to the RNA export adaptor, ALYREF; a component of the Nuclear Exosome Targeting (NEXT) complex, RBM7; and the nuclear cap binding complex (CBC). Regardless of function, all tested factors interact with nascent RNA as it exits RNAPII. Moreover, we demonstrate that the two transesterification steps of pre-mRNA splicing temporally separate ALYREF and RBM7 binding to splicing intermediates, and that exon-exon junction density drives RNA 5'end binding of ALYREF. Finally, we identify underappreciated steps in snoRNA 3'end processing performed by RBM7. Altogether, our data provide a temporal view of RNA-protein interactions during the early phases of transcription.
    DOI:  https://doi.org/10.1038/s41467-023-36345-y
  5. Nat Commun. 2023 Feb 04. 14(1): 608
    An updated suite of viral vectors for in vivo calcium imaging using intracerebral and retro-orbital injections in male mice.
    Sverre Grødem, Ingeborg Nymoen, Guro Helén Vatne, Frederik Sebastian Rogge, Valgerður Björnsdóttir, Kristian Kinden Lensjø, Marianne Fyhn.
      Genetically encoded Ca2+ indicators (GECIs) are widely used to measure neural activity. Here, we explore the use of systemically administered PHP.eB AAVs for brain-wide expression of GECIs and compare the expression properties to intracerebrally injected AAVs in male mice. We show that systemic administration is a promising strategy for imaging neural activity. Next, we establish the use of EE-RR- (soma) and RPL10a (Ribo) soma-targeting peptides with the latest jGCaMP and show that EE-RR-tagged jGCaMP8 gives rise to strong expression but limited soma-targeting. In contrast, Ribo-tagged jGCaMP8 lacks neuropil signal, but the expression rate is reduced. To combat this, we modified the linker region of the Ribo-tag (RiboL1-). RiboL1-jGCaMP8 expresses faster than Ribo-jGCaMP8 but remains too dim for reliable use with systemic virus administration. However, intracerebral injections of the RiboL1-tagged jGCaMP8 constructs provide strong Ca2+ signals devoid of neuropil contamination, with remarkable labeling density.
    DOI:  https://doi.org/10.1038/s41467-023-36324-3
  6. Nat Commun. 2023 Feb 07. 14(1): 660
    Myristic acid as a checkpoint to regulate STING-dependent autophagy and interferon responses by promoting N-myristoylation.
    Mutian Jia, Yuanyuan Wang, Jie Wang, Danhui Qin, Mengge Wang, Li Chai, Yue Fu, Chunyuan Zhao, Chengjiang Gao, Jihui Jia, Wei Zhao.
      Stimulator of interferon gene (STING)-triggered autophagy is crucial for the host to eliminate invading pathogens and serves as a self-limiting mechanism of STING-induced interferon (IFN) responses. Thus, the mechanisms that ensure the beneficial effects of STING activation are of particular importance. Herein, we show that myristic acid, a type of long-chain saturated fatty acid (SFA), specifically attenuates cGAS-STING-induced IFN responses in macrophages, while enhancing STING-dependent autophagy. Myristic acid inhibits HSV-1 infection-induced innate antiviral immune responses and promotes HSV-1 replication in mice in vivo. Mechanistically, myristic acid enhances N-myristoylation of ARF1, a master regulator that controls STING membrane trafficking. Consequently, myristic acid facilitates STING activation-triggered autophagy degradation of the STING complex. Thus, our work identifies myristic acid as a metabolic checkpoint that contributes to immune homeostasis by balancing STING-dependent autophagy and IFN responses. This suggests that myristic acid and N-myristoylation are promising targets for the treatment of diseases caused by aberrant STING activation.
    DOI:  https://doi.org/10.1038/s41467-023-36332-3
  7. Nat Commun. 2023 Feb 09. 14(1): 700
    TRIM40 is a pathogenic driver of inflammatory bowel disease subverting intestinal barrier integrity.
    Sujin Kang, Jaekyung Kim, Areum Park, Minsoo Koh, Wonji Shin, Gayoung Park, Taeyun A Lee, Hyung Jin Kim, Heonjong Han, Yongbo Kim, Myung Kyung Choi, Jae Hyung Park, Eunhye Lee, Hyun-Soo Cho, Hyun Woo Park, Jae Hee Cheon, Sungwook Lee, Boyoun Park.
      The cortical actin cytoskeleton plays a critical role in maintaining intestinal epithelial integrity, and the loss of this architecture leads to chronic inflammation, as seen in inflammatory bowel disease (IBD). However, the exact mechanisms underlying aberrant actin remodeling in pathological states remain largely unknown. Here, we show that a subset of patients with IBD exhibits substantially higher levels of tripartite motif-containing protein 40 (TRIM40), a gene that is hardly detectable in healthy individuals. TRIM40 is an E3 ligase that directly targets Rho-associated coiled-coil-containing protein kinase 1 (ROCK1), an essential kinase involved in promoting cell-cell junctions, markedly decreasing the phosphorylation of key signaling factors critical for cortical actin formation and stabilization. This causes failure of the epithelial barrier function, thereby promoting a long-lived inflammatory response. A mutant TRIM40 lacking the RING, B-box, or C-terminal domains has impaired ability to accelerate ROCK1 degradation-driven cortical actin disruption. Accordingly, Trim40-deficient male mice are highly resistant to dextran sulfate sodium (DSS)-induced colitis. Our findings highlight that aberrant upregulation of TRIM40, which is epigenetically silenced under healthy conditions, drives IBD by subverting cortical actin formation and exacerbating epithelial barrier dysfunction.
    DOI:  https://doi.org/10.1038/s41467-023-36424-0
  8. Nat Commun. 2023 Feb 09. 14(1): 551
    Splicing factor SRSF1 deficiency in the liver triggers NASH-like pathology and cell death.
    Waqar Arif, Bhoomika Mathur, Michael F Saikali, Ullas V Chembazhi, Katelyn Toohill, You Jin Song, Qinyu Hao, Saman Karimi, Steven M Blue, Brian A Yee, Eric L Van Nostrand, Sushant Bangru, Grace Guzman, Gene W Yeo, Kannanganattu V Prasanth, Sayeepriyadarshini Anakk, Carolyn L Cummins, Auinash Kalsotra.
      Regulation of RNA processing contributes profoundly to tissue development and physiology. Here, we report that serine-arginine-rich splicing factor 1 (SRSF1) is essential for hepatocyte function and survival. Although SRSF1 is mainly known for its many roles in mRNA metabolism, it is also crucial for maintaining genome stability. We show that acute liver damage in the setting of targeted SRSF1 deletion in mice is associated with the excessive formation of deleterious RNA-DNA hybrids (R-loops), which induce DNA damage. Combining hepatocyte-specific transcriptome, proteome, and RNA binding analyses, we demonstrate that widespread genotoxic stress following SRSF1 depletion results in global inhibition of mRNA transcription and protein synthesis, leading to impaired metabolism and trafficking of lipids. Lipid accumulation in SRSF1-deficient hepatocytes is followed by necroptotic cell death, inflammation, and fibrosis, resulting in NASH-like liver pathology. Importantly, SRSF1-depleted human liver cancer cells recapitulate this pathogenesis, illustrating a conserved and fundamental role for SRSF1 in preserving genome integrity and tissue homeostasis. Thus, our study uncovers how the accumulation of detrimental R-loops impedes hepatocellular gene expression, triggering metabolic derangements and liver damage.
    DOI:  https://doi.org/10.1038/s41467-023-35932-3
  9. Sci Immunol. 2023 Feb 17. 8(80): eabq5204
    Human IL-23 is essential for IFN-γ-dependent immunity to mycobacteria.
    Quentin Philippot, Masato Ogishi, Jonathan Bohlen, Julia Puchan, Andrés Augusto Arias, Tina Nguyen, Marta Martin-Fernandez, Clement Conil, Darawan Rinchai, Mana Momenilandi, Seyed Alireza Mahdaviani, Mohammad Keramatipour, Jérémie Rosain, Rui Yang, Taushif Khan, Anna-Lena Neehus, Marie Materna, Ji Eun Han, Jessica Peel, Federico Mele, Marc Weisshaar, Sandra Jovic, Paul Bastard, Romain Lévy, Tom Le Voyer, Peng Zhang, Majistor Raj Luxman Maglorius Renkilaraj, Carlos A Arango-Franco, Simon Pelham, Yoann Seeleuthner, Mathieu Pochon, Manar Mahmoud Ahmad Ata, Fatima Al Ali, Mélanie Migaud, Camille Soudée, Tatiana Kochetkov, Anne Molitor, Raphael Carapito, Seiamak Bahram, Bertrand Boisson, Claire Fieschi, Davood Mansouri, Nico Marr, Satoshi Okada, Mohammad Shahrooei, Nima Parvaneh, Zahra Chavoshzadeh, Aurélie Cobat, Dusan Bogunovic, Laurent Abel, Stuart G Tangye, Cindy S Ma, Vivien Béziat, Federica Sallusto, Stéphanie Boisson-Dupuis, Jacinta Bustamante, Jean-Laurent Casanova, Anne Puel.
      Patients with autosomal recessive (AR) IL-12p40 or IL-12Rβ1 deficiency display Mendelian susceptibility to mycobacterial disease (MSMD) due to impaired IFN-γ production and, less commonly, chronic mucocutaneous candidiasis (CMC) due to impaired IL-17A/F production. We report six patients from four kindreds with AR IL-23R deficiency. These patients are homozygous for one of four different loss-of-function IL23R variants. All six patients have a history of MSMD, but only two suffered from CMC. We show that IL-23 induces IL-17A only in MAIT cells, possibly contributing to the incomplete penetrance of CMC in patients unresponsive to IL-23. By contrast, IL-23 is required for both baseline and Mycobacterium-inducible IFN-γ immunity in both Vδ2+ γδ T and MAIT cells, probably contributing to the higher penetrance of MSMD in these patients. Human IL-23 appears to contribute to IL-17A/F-dependent immunity to Candida in a single lymphocyte subset but is required for IFN-γ-dependent immunity to Mycobacterium in at least two lymphocyte subsets.
    DOI:  https://doi.org/10.1126/sciimmunol.abq5204
  10. Nat Commun. 2023 Feb 10. 14(1): 741
    H2B ubiquitination recruits FACT to maintain a stable altered nucleosome state for transcriptional activation.
    Anfeng Luo, Jingwei Kong, Jun Chen, Xue Xiao, Jie Lan, Xiaorong Li, Cuifang Liu, Peng-Ye Wang, Guohong Li, Wei Li, Ping Chen.
      Histone H2B mono-ubiquitination at lysine 120 (ubH2B) has been found to regulate transcriptional elongation by collaborating with the histone chaperone FACT (Facilitates Chromatin Transcription) and plays essential roles in chromatin-based transcriptional processes. However, the mechanism of how ubH2B directly collaborates with FACT at the nucleosome level still remains elusive. In this study, we demonstrate that ubH2B impairs the mechanical stability of the nucleosome and helps to recruit FACT by enhancing the binding of FACT on the nucleosome. FACT prefers to bind and deposit H2A-ubH2B dimers to form an intact nucleosome. Strikingly, the preferable binding of FACT on ubH2B-nucleosome greatly enhances nucleosome stability and maintains its integrity. The stable altered nucleosome state obtained by ubH2B and FACT provides a key platform for gene transcription, as revealed by genome-wide and time-course ChIP-qPCR analyses. Our findings provide mechanistic insights of how ubH2B directly collaborates with FACT to regulate nucleosome dynamics for gene transcription.
    DOI:  https://doi.org/10.1038/s41467-023-36467-3
  11. Nat Commun. 2023 Feb 06. 14(1): 603
    LSD1/PRMT6-targeting gene therapy to attenuate androgen receptor toxic gain-of-function ameliorates spinobulbar muscular atrophy phenotypes in flies and mice.
    Ramachandran Prakasam, Angela Bonadiman, Roberta Andreotti, Emanuela Zuccaro, Davide Dalfovo, Caterina Marchioretti, Debasmita Tripathy, Gianluca Petris, Eric N Anderson, Alice Migazzi, Laura Tosatto, Anna Cereseto, Elena Battaglioli, Gianni Sorarù, Wooi Fang Lim, Carlo Rinaldi, Fabio Sambataro, Naemeh Pourshafie, Christopher Grunseich, Alessandro Romanel, Udai Bhan Pandey, Andrea Contestabile, Giuseppe Ronzitti, Manuela Basso, Maria Pennuto.
      Spinobulbar muscular atrophy (SBMA) is caused by CAG expansions in the androgen receptor gene. Androgen binding to polyQ-expanded androgen receptor triggers SBMA through a combination of toxic gain-of-function and loss-of-function mechanisms. Leveraging cell lines, mice, and patient-derived specimens, we show that androgen receptor co-regulators lysine-specific demethylase 1 (LSD1) and protein arginine methyltransferase 6 (PRMT6) are overexpressed in an androgen-dependent manner specifically in the skeletal muscle of SBMA patients and mice. LSD1 and PRMT6 cooperatively and synergistically transactivate androgen receptor, and their effect is enhanced by expanded polyQ. Pharmacological and genetic silencing of LSD1 and PRMT6 attenuates polyQ-expanded androgen receptor transactivation in SBMA cells and suppresses toxicity in SBMA flies, and a preclinical approach based on miRNA-mediated silencing of LSD1 and PRMT6 attenuates disease manifestations in SBMA mice. These observations suggest that targeting overexpressed co-regulators can attenuate androgen receptor toxic gain-of-function without exacerbating loss-of-function, highlighting a potential therapeutic strategy for patients with SBMA.
    DOI:  https://doi.org/10.1038/s41467-023-36186-9
  12. Nat Commun. 2023 Feb 09. 14(1): 709
    Adipocyte-derived extracellular vesicles increase insulin secretion through transport of insulinotropic protein cargo.
    Konxhe Kulaj, Alexandra Harger, Michaela Bauer, Özüm S Caliskan, Tilak Kumar Gupta, Dapi Menglin Chiang, Edward Milbank, Josefine Reber, Angelos Karlas, Petra Kotzbeck, David N Sailer, Francesco Volta, Dominik Lutter, Sneha Prakash, Juliane Merl-Pham, Vasilis Ntziachristos, Hans Hauner, Michael W Pfaffl, Matthias H Tschöp, Timo D Müller, Stefanie M Hauck, Benjamin D Engel, Jantje M Gerdes, Paul T Pfluger, Natalie Krahmer, Kerstin Stemmer.
      Adipocyte-derived extracellular vesicles (AdEVs) are membranous nanoparticles that convey communication from adipose tissue to other organs. Here, to delineate their role as messengers with glucoregulatory nature, we paired fluorescence AdEV-tracing and SILAC-labeling with (phospho)proteomics, and revealed that AdEVs transfer functional insulinotropic protein cargo into pancreatic β-cells. Upon transfer, AdEV proteins were subjects for phosphorylation, augmented insulinotropic GPCR/cAMP/PKA signaling by increasing total protein abundances and phosphosite dynamics, and ultimately enhanced 1st-phase glucose-stimulated insulin secretion (GSIS) in murine islets. Notably, insulinotropic effects were restricted to AdEVs isolated from obese and insulin resistant, but not lean mice, which was consistent with differential protein loads and AdEV luminal morphologies. Likewise, in vivo pre-treatment with AdEVs from obese but not lean mice amplified insulin secretion and glucose tolerance in mice. This data suggests that secreted AdEVs can inform pancreatic β-cells about insulin resistance in adipose tissue in order to amplify GSIS in times of increased insulin demand.
    DOI:  https://doi.org/10.1038/s41467-023-36148-1
  13. Sci Transl Med. 2023 Feb 08. 15(682): eadc9653
    Hepatocytes demarcated by EphB2 contribute to the progression of nonalcoholic steatohepatitis.
    Yang Xiao, Kirill Batmanov, Wenxiang Hu, Kun Zhu, Alexander Y Tom, Dongyin Guan, Chunjie Jiang, Lan Cheng, Sam J McCright, Eric C Yang, Matthew R Lanza, Yifan Liu, David A Hill, Mitchell A Lazar.
      Current therapeutic strategies for treating nonalcoholic steatohepatitis (NASH) have failed to alleviate liver fibrosis, which is a devastating feature leading to hepatic dysfunction. Here, we integrated single-nucleus transcriptomics and epigenomics to characterize all major liver cell types during NASH development in mice and humans. The bifurcation of hepatocyte trajectory with NASH progression was conserved between mice and humans. At the nonalcoholic fatty liver (NAFL) stage, hepatocytes exhibited metabolic adaptation, whereas at the NASH stage, a subset of hepatocytes was enriched for the signatures of cell adhesion and migration, which were mainly demarcated by receptor tyrosine kinase ephrin type B receptor 2 (EphB2). EphB2, acting as a downstream effector of Notch signaling in hepatocytes, was sufficient to induce cell-autonomous inflammation. Knockdown of Ephb2 in hepatocytes ameliorated inflammation and fibrosis in a mouse model of NASH. Thus, EphB2-expressing hepatocytes contribute to NASH progression and may serve as a potential therapeutic target.
    DOI:  https://doi.org/10.1126/scitranslmed.adc9653
  14. Nat Commun. 2023 Feb 11. 14(1): 766
    Lipid-droplet associated mitochondria promote fatty-acid oxidation through a distinct bioenergetic pattern in male Wistar rats.
    Noble Kumar Talari, Ushodaya Mattam, Niroj Kumar Meher, Arun Kumar Paripati, Kalyankar Mahadev, Thanuja Krishnamoorthy, Naresh Babu V Sepuri.
      Mitochondria empower the liver to regulate lipid homeostasis by enabling fatty acid oxidation during starvation and lipogenesis during nutrient-rich conditions. It is unknown if mitochondria can seamlessly regulate these two distinct processes or if two discrete populations of mitochondria achieve these two functions in the liver. For the first time in the liver, we report the isolation of two distinct populations of mitochondria from male Wistar rats on an ad-libitum diet: cytoplasmic mitochondria and lipid droplet-associated mitochondria. Our studies show that while lipid droplet mitochondria exhibit higher fatty acid oxidation and are marked by enhanced levels of pACC2, MFN2, and CPT1 activity, cytoplasmic mitochondria are associated with higher respiration capacity. Notably, lipid droplet-associated mitochondria isolated from a non-alcoholic fatty liver disease (NAFLD) rat model are compromised for fatty acid oxidation. We demonstrate the importance of functional segregation of mitochondria as any aberration in lipid droplet-associated mitochondria may lead to NAFLD.
    DOI:  https://doi.org/10.1038/s41467-023-36432-0
  15. Cell. 2023 Feb 06. pii: S0092-8674(23)00004-1. [Epub ahead of print]
    Extending genetic risk for Alzheimer's disease from host to holobiont.
    Sabeen A Kazmi, Elaine Y Hsiao.
      The gut microbiota is implicated in risk for Alzheimer's disease (AD). A study in Science reports that depleting gut bacteria in mice with genetic risk for AD reduces neuropathology in a sex-dependent manner. This is reversed by administering short-chain fatty acids, suggesting that specific bacterial metabolites increase susceptibility to AD.
    DOI:  https://doi.org/10.1016/j.cell.2023.01.004
  16. Nat Commun. 2023 Feb 09. 14(1): 632
    ZBTB12 is a molecular barrier to dedifferentiation in human pluripotent stem cells.
    Dasol Han, Guojing Liu, Yujeong Oh, Seyoun Oh, Seungbok Yang, Lori Mandjikian, Neha Rani, Maria C Almeida, Kenneth S Kosik, Jiwon Jang.
      Development is generally viewed as one-way traffic of cell state transition from primitive to developmentally advanced states. However, molecular mechanisms that ensure the unidirectional transition of cell fates remain largely unknown. Through exact transcription start site mapping, we report an evolutionarily conserved BTB domain-containing zinc finger protein, ZBTB12, as a molecular barrier for dedifferentiation of human pluripotent stem cells (hPSCs). Single-cell RNA sequencing reveals that ZBTB12 is essential for three germ layer differentiation by blocking hPSC dedifferentiation. Mechanistically, ZBTB12 fine-tunes the expression of human endogenous retrovirus H (HERVH), a primate-specific retrotransposon, and targets specific transcripts that utilize HERVH as a regulatory element. In particular, the downregulation of HERVH-overlapping long non-coding RNAs (lncRNAs) by ZBTB12 is necessary for a successful exit from a pluripotent state and lineage derivation. Overall, we identify ZBTB12 as a molecular barrier that safeguards the unidirectional transition of metastable stem cell fates toward developmentally advanced states.
    DOI:  https://doi.org/10.1038/s41467-023-36178-9
  17. Science. 2023 Feb 10. 379(6632): 614
    What no one told me.
    Halley Mastro.
      
    DOI:  https://doi.org/10.1126/science.adh0049
  18. Nat Commun. 2023 Feb 10. 14(1): 749
    Application of high-throughput single-nucleus DNA sequencing in pancreatic cancer.
    Haochen Zhang, Elias-Ramzey Karnoub, Shigeaki Umeda, Ronan Chaligné, Ignas Masilionis, Caitlin A McIntyre, Palash Sashittal, Akimasa Hayashi, Amanda Zucker, Katelyn Mullen, Jungeui Hong, Alvin Makohon-Moore, Christine A Iacobuzio-Donahue.
      Despite insights gained by bulk DNA sequencing of cancer it remains challenging to resolve the admixture of normal and tumor cells, and/or of distinct tumor subclones; high-throughput single-cell DNA sequencing circumvents these and brings cancer genomic studies to higher resolution. However, its application has been limited to liquid tumors or a small batch of solid tumors, mainly because of the lack of a scalable workflow to process solid tumor samples. Here we optimize a highly automated nuclei extraction workflow that achieves fast and reliable targeted single-nucleus DNA library preparation of 38 samples from 16 pancreatic ductal adenocarcinoma patients, with an average library yield per sample of 2867 single nuclei. We demonstrate that this workflow not only performs well using low cellularity or low tumor purity samples but reveals genomic evolution patterns of pancreatic ductal adenocarcinoma as well.
    DOI:  https://doi.org/10.1038/s41467-023-36344-z
  19. Cell Rep. 2023 Feb 10. pii: S2211-1247(23)00117-1. [Epub ahead of print]42(2): 112106
    DRAK2 contributes to type 1 diabetes by negatively regulating IL-2 sensitivity to alter regulatory T cell development.
    Alexandra H Mandarano, Tarsha L Harris, Blaine M Creasy, Marie Wehenkel, Marygrace Duggar, Benjamin A Wilander, Ashutosh Mishra, Jeremy Chase Crawford, Sarah A Mullen, Katherine M Williams, Meenu Pillai, Anthony A High, Maureen A McGargill.
      Drak2-deficient (Drak2-/-) mice are resistant to multiple models of autoimmunity yet effectively eliminate pathogens and tumors. Thus, DRAK2 represents a potential target to treat autoimmune diseases. However, the mechanisms by which DRAK2 contributes to autoimmunity, particularly type 1 diabetes (T1D), remain unresolved. Here, we demonstrate that resistance to T1D in non-obese diabetic (NOD) mice is due to the absence of Drak2 in T cells and requires the presence of regulatory T cells (Tregs). Contrary to previous hypotheses, we show that DRAK2 does not limit TCR signaling. Rather, DRAK2 regulates IL-2 signaling by inhibiting STAT5A phosphorylation. We further demonstrate that enhanced sensitivity to IL-2 in the absence of Drak2 augments thymic Treg development. Overall, our data indicate that DRAK2 contributes to autoimmunity in multiple ways by regulating thymic Treg development and by impacting the sensitivity of conventional T cells to Treg-mediated suppression.
    Keywords:  CP: Immunology; IL-2; T cells; regulatory T cells; thymus; tolerance; type 1 diabetes
    DOI:  https://doi.org/10.1016/j.celrep.2023.112106
  20. Cell. 2023 Feb 02. pii: S0092-8674(23)00007-7. [Epub ahead of print]
    Recycling of modified H2A-H2B provides short-term memory of chromatin states.
    Valentin Flury, Nazaret Reverón-Gómez, Nicolas Alcaraz, Kathleen R Stewart-Morgan, Alice Wenger, Robert J Klose, Anja Groth.
      Chromatin landscapes are disrupted during DNA replication and must be restored faithfully to maintain genome regulation and cell identity. The histone H3-H4 modification landscape is restored by parental histone recycling and modification of new histones. How DNA replication impacts on histone H2A-H2B is currently unknown. Here, we measure H2A-H2B modifications and H2A.Z during DNA replication and across the cell cycle using quantitative genomics. We show that H2AK119ub1, H2BK120ub1, and H2A.Z are recycled accurately during DNA replication. Modified H2A-H2B are segregated symmetrically to daughter strands via POLA1 on the lagging strand, but independent of H3-H4 recycling. Post-replication, H2A-H2B modification and variant landscapes are quickly restored, and H2AK119ub1 guides accurate restoration of H3K27me3. This work reveals epigenetic transmission of parental H2A-H2B during DNA replication and identifies cross talk between H3-H4 and H2A-H2B modifications in epigenome propagation. We propose that rapid short-term memory of recycled H2A-H2B modifications facilitates restoration of stable H3-H4 chromatin states.
    Keywords:  DNA replication; H2A; H2A.Z; H2B; chromatin; histone PTM cross talk; histone recycling; polycomb; post-translational modifications; ubiquitination
    DOI:  https://doi.org/10.1016/j.cell.2023.01.007
  21. Nat Commun. 2023 Feb 09. 14(1): 732
    A conformational switch in clathrin light chain regulates lattice structure and endocytosis at the plasma membrane of mammalian cells.
    Kazuki Obashi, Kem A Sochacki, Marie-Paule Strub, Justin W Taraska.
      Conformational changes in endocytic proteins are regulators of clathrin-mediated endocytosis. Three clathrin heavy chains associated with clathrin light chains (CLC) assemble into triskelia that link into a geometric lattice that curves to drive endocytosis. Structural changes in CLC have been shown to regulate triskelia assembly in solution, yet the nature of these changes, and their effects on lattice growth, curvature, and endocytosis in cells are unknown. Here, we develop a new correlative fluorescence resonance energy transfer (FRET) and platinum replica electron microscopy method, named FRET-CLEM. With FRET-CLEM, we measure conformational changes in clathrin at thousands of individual morphologically distinct clathrin-coated structures. We discover that the N-terminus of CLC repositions away from the plasma membrane and triskelia vertex as coats curve. Preventing this conformational switch with chemical tools increases lattice sizes and inhibits endocytosis. Thus, a specific conformational switch in the light chain regulates lattice curvature and endocytosis in mammalian cells.
    DOI:  https://doi.org/10.1038/s41467-023-36304-7
  22. Trends Immunol. 2023 Feb 02. pii: S1471-4906(23)00017-0. [Epub ahead of print]
    Oxidized mitochondrial DNA: a protective signal gone awry.
    Hongxu Xian, Michael Karin.
      Despite the emergence of mitochondria as key regulators of innate immunity, the mechanisms underlying the generation and release of immunostimulatory alarmins by stressed mitochondria remains nebulous. We propose that the major mitochondrial alarmin in myeloid cells is oxidized mitochondrial DNA (Ox-mtDNA). Fragmented Ox-mtDNA enters the cytosol where it activates the NLRP3 inflammasome and generates IL-1β, IL-18, and cGAS-STING to induce type I interferons and interferon-stimulated genes. Inflammasome activation further enables the circulatory release of Ox-mtDNA by opening gasdermin D pores. We summarize new data showing that, in addition to being an autoimmune disease biomarker, Ox-mtDNA converts beneficial transient inflammation into long-lasting immunopathology. We discuss how Ox-mtDNA induces short- and long-term immune activation, and highlight its homeostatic and immunopathogenic functions.
    Keywords:  NLRP3 inflammasome; Ox-mtDNA; autoimmunity; cGAS–STING; cell-free DNA; immunopathology; inflammation; stressed mitochondria
    DOI:  https://doi.org/10.1016/j.it.2023.01.006
  23. Nat Commun. 2023 Feb 06. 14(1): 602
    Defective excitation-contraction coupling and mitochondrial respiration precede mitochondrial Ca2+ accumulation in spinobulbar muscular atrophy skeletal muscle.
    Caterina Marchioretti, Giulia Zanetti, Marco Pirazzini, Gaia Gherardi, Leonardo Nogara, Roberta Andreotti, Paolo Martini, Lorenzo Marcucci, Marta Canato, Samir R Nath, Emanuela Zuccaro, Mathilde Chivet, Cristina Mammucari, Marco Pacifici, Anna Raffaello, Rosario Rizzuto, Andrea Mattarei, Maria A Desbats, Leonardo Salviati, Aram Megighian, Gianni Sorarù, Elena Pegoraro, Elisa Belluzzi, Assunta Pozzuoli, Carlo Biz, Pietro Ruggieri, Chiara Romualdi, Andrew P Lieberman, Gopal J Babu, Marco Sandri, Bert Blaauw, Manuela Basso, Maria Pennuto.
      Polyglutamine expansion in the androgen receptor (AR) causes spinobulbar muscular atrophy (SBMA). Skeletal muscle is a primary site of toxicity; however, the current understanding of the early pathological processes that occur and how they unfold during disease progression remains limited. Using transgenic and knock-in mice and patient-derived muscle biopsies, we show that SBMA mice in the presymptomatic stage develop a respiratory defect matching defective expression of genes involved in excitation-contraction coupling (ECC), altered contraction dynamics, and increased fatigue. These processes are followed by stimulus-dependent accumulation of calcium into mitochondria and structural disorganization of the muscle triads. Deregulation of expression of ECC genes is concomitant with sexual maturity and androgen raise in the serum. Consistent with the androgen-dependent nature of these alterations, surgical castration and AR silencing alleviate the early and late pathological processes. These observations show that ECC deregulation and defective mitochondrial respiration are early but reversible events followed by altered muscle force, calcium dyshomeostasis, and dismantling of triad structure.
    DOI:  https://doi.org/10.1038/s41467-023-36185-w
  24. Nat Commun. 2023 Feb 09. 14(1): 717
    Light-driven biological actuators to probe the rheology of 3D microtissues.
    Adrien Méry, Artur Ruppel, Jean Revilloud, Martial Balland, Giovanni Cappello, Thomas Boudou.
      The mechanical properties of biological tissues are key to their physical integrity and function. Although external loading or biochemical treatments allow the estimation of these properties globally, it remains difficult to assess how such external stimuli compare with cell-generated contractions. Here we engineer microtissues composed of optogenetically-modified fibroblasts encapsulated within collagen. Using light to control the activity of RhoA, a major regulator of cellular contractility, we induce local contractions within microtissues, while monitoring microtissue stress and strain. We investigate the regulation of these local contractions and their spatio-temporal distribution. We demonstrate the potential of our technique for quantifying tissue elasticity and strain propagation, before examining the possibility of using light to create and map local anisotropies in mechanically heterogeneous microtissues. Altogether, our results open an avenue to guide the formation of tissues while non-destructively charting their rheology in real time, using their own constituting cells as internal actuators.
    DOI:  https://doi.org/10.1038/s41467-023-36371-w
  25. Nat Commun. 2023 Feb 11. 14(1): 769
    Histone variant H2A.Z modulates nucleosome dynamics to promote DNA accessibility.
    Shuxiang Li, Tiejun Wei, Anna R Panchenko.
      Nucleosomes, containing histone variants H2A.Z, are important for gene transcription initiation and termination, chromosome segregation and DNA double-strand break repair, among other functions. However, the underlying mechanisms of how H2A.Z influences nucleosome stability, dynamics and DNA accessibility are not well understood, as experimental and computational evidence remains inconclusive. Our modeling efforts of human nucleosome stability and dynamics, along with comparisons with experimental data show that the incorporation of H2A.Z results in a substantial decrease of the energy barrier for DNA unwrapping. This leads to the spontaneous DNA unwrapping of about forty base pairs from both ends, nucleosome gapping and increased histone plasticity, which otherwise is not observed for canonical nucleosomes. We demonstrate that both N- and C-terminal tails of H2A.Z play major roles in these events, whereas the H3.3 variant exerts a negligible impact in modulating the DNA end unwrapping. In summary, our results indicate that H2A.Z deposition makes nucleosomes more mobile and DNA more accessible to transcriptional machinery and other chromatin components.
    DOI:  https://doi.org/10.1038/s41467-023-36465-5
  26. Nat Commun. 2023 Feb 06. 14(1): 642
    Microbiota-derived acetate enhances host antiviral response via NLRP3.
    Junling Niu, Mengmeng Cui, Xin Yang, Juan Li, Yuhui Yao, Qiuhong Guo, Ailing Lu, Xiaopeng Qi, Dongming Zhou, Chenhong Zhang, Liping Zhao, Guangxun Meng.
      Pathogenic viral infections represent a major challenge to human health. Host immune responses to respiratory viruses are closely associated with microbiome and metabolism via the gut-lung axis. It has been known that host defense against influenza A virus (IAV) involves activation of the NLRP3 inflammasome, however, mechanisms behind the protective function of NLRP3 are not fully known. Here we show that an isolated bacterial strain, Bifidobacterium pseudolongum NjM1, enriched in the gut microbiota of Nlrp3-/- mice, protects wild-type but not Nlrp3 deficient mice against IAV infection. This effect depends on the enhanced production of type I interferon (IFN-I) mediated by NjM1-derived acetate. Application of exogenous acetate reproduces the protective effect of NjM1. Mechanistically, NLRP3 bridges GPR43 and MAVS, and promotes the oligomerization and signalling of MAVS; while acetate enhances MAVS aggregation upon GPR43 engagement, leading to elevated IFN-I production. Thus, our data support a model of NLRP3 mediating enhanced induction of IFN-I via acetate-producing bacterium and suggest that the acetate-GPR43-NLRP3-MAVS-IFN-I signalling axis is a potential therapeutic target against respiratory viral infections.
    DOI:  https://doi.org/10.1038/s41467-023-36323-4
  27. Cell. 2023 Feb 03. pii: S0092-8674(23)00042-9. [Epub ahead of print]
    Cryo-EM structure of the RADAR supramolecular anti-phage defense complex.
    Brianna Duncan-Lowey, Nitzan Tal, Alex G Johnson, Shaun Rawson, Megan L Mayer, Shany Doron, Adi Millman, Sarah Melamed, Taya Fedorenko, Assaf Kacen, Alexander Brandis, Tevie Mehlman, Gil Amitai, Rotem Sorek, Philip J Kranzusch.
      RADAR is a two-protein bacterial defense system that was reported to defend against phage by "editing" messenger RNA. Here, we determine cryo-EM structures of the RADAR defense complex, revealing RdrA as a heptameric, two-layered AAA+ ATPase and RdrB as a dodecameric, hollow complex with twelve surface-exposed deaminase active sites. RdrA and RdrB join to form a giant assembly up to 10 MDa, with RdrA docked as a funnel over the RdrB active site. Surprisingly, our structures reveal an RdrB active site that targets mononucleotides. We show that RdrB catalyzes ATP-to-ITP conversion in vitro and induces the massive accumulation of inosine mononucleotides during phage infection in vivo, limiting phage replication. Our results define ATP mononucleotide deamination as a determinant of RADAR immunity and reveal supramolecular assembly of a nucleotide-modifying machine as a mechanism of anti-phage defense.
    Keywords:  adenosine deaminase; anti-phage immunity; phage
    DOI:  https://doi.org/10.1016/j.cell.2023.01.012
  28. Nat Commun. 2023 Feb 08. 14(1): 679
    Cartography of Genomic Interactions Enables Deep Analysis of Single-Cell Expression Data.
    Md Tauhidul Islam, Lei Xing.
      Remarkable advances in single cell genomics have presented unique challenges and opportunities for interrogating a wealth of biomedical inquiries. High dimensional genomic data are inherently complex because of intertwined relationships among the genes. Existing methods, including emerging deep learning-based approaches, do not consider the underlying biological characteristics during data processing, which greatly compromises the performance of data analysis and hinders the maximal utilization of state-of-the-art genomic techniques. In this work, we develop an entropy-based cartography strategy to contrive the high dimensional gene expression data into a configured image format, referred to as genomap, with explicit integration of the genomic interactions. This unique cartography casts the gene-gene interactions into the spatial configuration of genomaps and enables us to extract the deep genomic interaction features and discover underlying discriminative patterns of the data. We show that, for a wide variety of applications (cell clustering and recognition, gene signature extraction, single cell data integration, cellular trajectory analysis, dimensionality reduction, and visualization), the proposed approach drastically improves the accuracies of data analyses as compared to the state-of-the-art techniques.
    DOI:  https://doi.org/10.1038/s41467-023-36383-6
  29. Nat Immunol. 2023 Feb 09.
    The endogenous repertoire harbors self-reactive CD4+ T cell clones that adopt a follicular helper T cell-like phenotype at steady state.
    Victoria Lee, Donald M Rodriguez, Nicole K Ganci, Sharon Zeng, Junting Ai, Jaime L Chao, Matthew T Walker, Christine H Miller, David E J Klawon, Mary H Schoenbach, Domenick E Kennedy, Mark Maienschein-Cline, Nicholas D Socci, Marcus R Clark, Peter A Savage.
      The T cell repertoire of healthy mice and humans harbors self-reactive CD4+ conventional T (Tconv) cells capable of inducing autoimmunity. Using T cell receptor profiling paired with in vivo clonal analysis of T cell differentiation, we identified Tconv cell clones that are recurrently enriched in non-lymphoid organs following ablation of Foxp3+ regulatory T (Treg) cells. A subset of these clones was highly proliferative in the lymphoid organs at steady state and exhibited overt reactivity to self-ligands displayed by dendritic cells, yet were not purged by clonal deletion. These clones spontaneously adopted numerous hallmarks of follicular helper T (TFH) cells, including expression of Bcl6 and PD-1, exhibited an elevated propensity to localize within B cell follicles at steady state, and produced interferon-γ in non-lymphoid organs following sustained Treg cell depletion. Our work identifies a naturally occurring population of self-reactive TFH-like cells and delineates a previously unappreciated fate for self-specific Tconv cells.
    DOI:  https://doi.org/10.1038/s41590-023-01425-0
  30. Commun Biol. 2023 02 08. 6(1): 157
    Melatonin protects against cadmium-induced oxidative stress via mitochondrial STAT3 signaling in human prostate stromal cells.
    Moonjung Hyun, Hyejin Kim, Jehein Kim, Juhong Lee, Ho Jeong Lee, Laxmi Rathor, Jeremy Meier, Andrew Larner, Seon Min Lee, Yeongyu Moon, Jungil Choi, Sung Min Han, Jeong-Doo Heo.
      Melatonin protects against Cadmium (Cd)-induced toxicity, a ubiquitous environmental toxicant that causes adverse health effects by increasing reactive oxygen species (ROS) production and mitochondrial dysfunction. However, the underlying mechanism remains unclear. Here, we demonstrate that Cd exposure reduces the levels of mitochondrially-localized signal transducer and activator of transcription 3 (mitoSTAT3) using human prostate stromal cells and mouse embryonic fibroblasts. Melatonin enhances mitoSTAT3 abundance following Cd exposure, which is required to attenuate ROS damage, mitochondrial dysfunction, and cell death caused by Cd exposure. Moreover, melatonin increases mitochondrial levels of GRIM-19, an electron transport chain component that mediates STAT3 import into mitochondria, which are downregulated by Cd. In vivo, melatonin reverses the reduced size of mouse prostate tissue and levels of mitoSTAT3 and GRIM-19 induced by Cd exposure. Together, these data suggest that melatonin regulates mitoSTAT3 function to prevent Cd-induced cytotoxicity and could preserve mitochondrial function during Cd-induced stress.
    DOI:  https://doi.org/10.1038/s42003-023-04533-7
  31. Sci Transl Med. 2023 Feb 08. 15(682): eadd6373
    Peanut allergen inhibition prevents anaphylaxis in a humanized mouse model.
    Nada S Alakhras, Jaeho Shin, Scott A Smith, Anthony L Sinn, Wenwu Zhang, Gyoyeon Hwang, Jenna Sjoerdsma, Emily K Bromley, Karen E Pollok, Basar Bilgicer, Mark H Kaplan.
      Peanut-induced allergy is an immunoglobulin E (IgE)-mediated type I hypersensitivity reaction that manifests symptoms ranging from local edema to life-threatening anaphylaxis. Although there are treatments for symptoms in patients with allergies resulting from allergen exposure, there are few preventive therapies other than strict dietary avoidance or oral immunotherapy, neither of which are successful in all patients. We have previously designed a covalent heterobivalent inhibitor (cHBI) that binds in an allergen-specific manner as a preventive for allergic reactions. Building on previous in vitro testing, here, we developed a humanized mouse model to test cHBI efficacy in vivo. Nonobese diabetic-severe combined immunodeficient γc-deficient mice expressing transgenes for human stem cell factor, granulocyte-macrophage colony-stimulating factor, and interleukin-3 developed mature functional human mast cells in multiple tissues and displayed robust anaphylactic reactions when passively sensitized with patient-derived IgE monoclonal antibodies specific for peanut Arachis hypogaea 2 (Ara h 2). The allergic response in humanized mice was IgE dose dependent and was mediated by human mast cells. Using this humanized mouse model, we showed that cHBI prevented allergic reactions for more than 2 weeks when administered before allergen exposure. cHBI also prevented fatal anaphylaxis and attenuated allergic reactions when administered shortly after the onset of symptoms. cHBI impaired mast cell degranulation in vivo in an allergen-specific manner. cHBI rescued the mice from lethal anaphylactic responses during oral Ara h 2 allergen-induced anaphylaxis. Together, these findings suggest that cHBI has the potential to be an effective preventative for peanut-specific allergic responses in patients.
    DOI:  https://doi.org/10.1126/scitranslmed.add6373
  32. Cell Genom. 2022 Nov 09. pii: 100191. [Epub ahead of print]2(11):
    A cis-regulatory lexicon of DNA motif combinations mediating cell-type-specific gene regulation.
    Laura K H Donohue, Margaret G Guo, Yang Zhao, Namyoung Jung, Rose T Bussat, Daniel S Kim, Poornima H Neela, Laura N Kellman, Omar S Garcia, Robin M Meyers, Russ B Altman, Paul A Khavari.
      Gene expression is controlled by transcription factors (TFs) that bind cognate DNA motif sequences in cis-regulatory elements (CREs). The combinations of DNA motifs acting within homeostasis and disease, however, are unclear. Gene expression, chromatin accessibility, TF footprinting, and H3K27ac-dependent DNA looping data were generated and a random-forest-based model was applied to identify 7,531 cell-type-specific cis-regulatory modules (CRMs) across 15 diploid human cell types. A co-enrichment framework within CRMs nominated 838 cell-type-specific, recurrent heterotypic DNA motif combinations (DMCs), which were functionally validated using massively parallel reporter assays. Cancer cells engaged DMCs linked to neoplasia-enabling processes operative in normal cells while also activating new DMCs only seen in the neoplastic state. This integrative approach identifies cell-type-specific cis-regulatory combinatorial DNA motifs in diverse normal and diseased human cells and represents a general framework for deciphering cis-regulatory sequence logic in gene regulation.
    DOI:  https://doi.org/10.1016/j.xgen.2022.100191
  33. Proc Natl Acad Sci U S A. 2023 Feb 14. 120(7): e2207909120
    Hippocampal-cortical coupling differentiates long-term memory processes.
    Prawesh Dahal, Onni J Rauhala, Dion Khodagholy, Jennifer N Gelinas.
      Reactivation of long-term memories enables experience-dependent strengthening, weakening, or updating of memory traces. Although coupling of hippocampal and cortical activity patterns facilitates initial memory consolidation, whether and how these patterns are involved in postreactivation memory processes are not known. Here, we monitored the hippocampal-cortical network as rats repetitively learned and retrieved spatial and nonspatial memories. We show that interactions between hippocampal sharp wave-ripples (SPW-R), cortical spindles (SPI), and cortical ripples (CXR) are jointly modulated in the absence of memory demand but independently recruited depending on the stage of memory and task type. Reconsolidation of memory after retrieval is associated with an increased and extended window of coupling between hippocampal SPW-Rs and CXRs compared to the initial consolidation. Hippocampal SPW-R and cortical spindle interactions are preferentially engaged during memory consolidation. These findings suggest that specific, time-limited patterns of oscillatory coupling can support the distinct memory processes required to flexibly manage long-term memories in a dynamic environment.
    Keywords:  consolidation; hippocampus; learning; memory; ripples
    DOI:  https://doi.org/10.1073/pnas.2207909120
  34. Nat Aging. 2022 May;2(5): 379-388
    Limited Proteolysis-Mass Spectrometry Reveals Aging-Associated Changes in Cerebrospinal Fluid Protein Abundances and Structures.
    Steven R Shuken, Jarod Rutledge, Tal Iram, Patricia Moran Losada, Edward N Wilson, Katrin I Andreasson, Ryan D Leib, Tony Wyss-Coray.
      Cerebrospinal fluid (CSF) proteins and their structures have been implicated repeatedly in aging and neurodegenerative diseases. Limited proteolysis-mass spectrometry (LiP-MS) is a method that enables proteome-wide screening for changes in both protein abundance and structure. To screen for novel aging-associated changes in the CSF proteome, we performed LiP-MS on CSF from young and old mice with a modified analysis pipeline. We found 38 protein groups change in abundance with aging, most dominantly immunoglobulins of the IgM subclass. We discovered six high-confidence candidates that appeared to change in structure with aging, of which Kng1, Itih2, Lp-PLA2, and 14-3-3 proteins have binding partners or proteoforms known previously to change in the brain with Alzheimer's disease. Intriguingly, using orthogonal validation by Western blot we found the LiP-MS hit Cd5l forms a covalent complex with IgM in mouse and human CSF whose abundance increases with aging. SOMAmer probe signals for all six LiP-MS hits in human CSF, especially 14-3-3 proteins, significantly associate with several clinical features relevant to cognitive function and neurodegeneration. Together, our findings show that LiP-MS can uncover age-related structural changes in CSF with relevance to neurodegeneration.
    DOI:  https://doi.org/10.1038/s43587-022-00196-x
  35. Nature. 2023 Feb 08.
    TET2 guards against unchecked BATF3-induced CAR T cell expansion.
    Nayan Jain, Zeguo Zhao, Judith Feucht, Richard Koche, Archana Iyer, Anton Dobrin, Jorge Mansilla-Soto, Julie Yang, Yingqian Zhan, Michael Lopez, Gertrude Gunset, Michel Sadelain.
      Further advances in cell engineering are needed to increase the efficacy of chimeric antigen receptor (CAR) and other T cell-based therapies1-5. As T cell differentiation and functional states are associated with distinct epigenetic profiles6,7, we hypothesized that epigenetic programming may provide a means to improve CAR T cell performance. Targeting the gene that encodes the epigenetic regulator ten-eleven translocation 2 (TET2)8 presents an interesting opportunity as its loss may enhance T cell memory9,10, albeit not cause malignancy9,11,12. Here we show that disruption of TET2 enhances T cell-mediated tumour rejection in leukaemia and prostate cancer models. However, loss of TET2 also enables antigen-independent CAR T cell clonal expansions that may eventually result in prominent systemic tissue infiltration. These clonal proliferations require biallelic TET2 disruption and sustained expression of the AP-1 factor BATF3 to drive a MYC-dependent proliferative program. This proliferative state is associated with reduced effector function that differs from both canonical T cell memory13,14 and exhaustion15,16 states, and is prone to the acquisition of secondary somatic mutations, establishing TET2 as a guardian against BATF3-induced CAR T cell proliferation and ensuing genomic instability. Our findings illustrate the potential of epigenetic programming to enhance T cell immunity but highlight the risk of unleashing unchecked proliferative responses.
    DOI:  https://doi.org/10.1038/s41586-022-05692-z
  36. Cell Metab. 2023 Feb 07. pii: S1550-4131(23)00003-7. [Epub ahead of print]35(2): 345-360.e7
    Mitochondrial fragmentation and donut formation enhance mitochondrial secretion to promote osteogenesis.
    Joonho Suh, Na-Kyung Kim, Wonn Shim, Seung-Hoon Lee, Hyo-Jeong Kim, Eunyoung Moon, Hiromi Sesaki, Jae Hyuck Jang, Jung-Eun Kim, Yun-Sil Lee.
      Mitochondrial components have been abundantly detected in bone matrix, implying that they are somehow transported extracellularly to regulate osteogenesis. Here, we demonstrate that mitochondria and mitochondrial-derived vesicles (MDVs) are secreted from mature osteoblasts to promote differentiation of osteoprogenitors. We show that osteogenic induction stimulates mitochondrial fragmentation, donut formation, and secretion of mitochondria through CD38/cADPR signaling. Enhancing mitochondrial fission and donut formation through Opa1 knockdown or Fis1 overexpression increases mitochondrial secretion and accelerates osteogenesis. We also show that mitochondrial fusion promoter M1, which induces Opa1 expression, impedes osteogenesis, whereas osteoblast-specific Opa1 deletion increases bone mass. We further demonstrate that secreted mitochondria and MDVs enhance bone regeneration in vivo. Our findings suggest that mitochondrial morphology in mature osteoblasts is adapted for extracellular secretion, and secreted mitochondria and MDVs are critical promoters of osteogenesis.
    Keywords:  FIS1; M1; OPA1; donut mitochondria; mitochondria; mitochondrial secretion; mitochondrial transplantation; mitochondrial-derived vesicles; osteoblasts; osteogenesis
    DOI:  https://doi.org/10.1016/j.cmet.2023.01.003
  37. Nat Commun. 2023 Feb 04. 14(1): 616
    Cell surface-bound La protein regulates the cell fusion stage of osteoclastogenesis.
    Jarred M Whitlock, Evgenia Leikina, Kamran Melikov, Luis Fernandez De Castro, Sandy Mattijssen, Richard J Maraia, Michael T Collins, Leonid V Chernomordik.
      Multinucleated osteoclasts, essential for skeletal remodeling in health and disease, are formed by the fusion of osteoclast precursors, where each fusion event raises their bone-resorbing activity. Here we show that the nuclear RNA chaperone, La protein has an additional function as an osteoclast fusion regulator. Monocyte-to-osteoclast differentiation starts with a drastic decrease in La levels. As fusion begins, La reappears as a low molecular weight species at the osteoclast surface, where it promotes fusion. La's role in promoting osteoclast fusion is independent of canonical La-RNA interactions and involves direct interactions between La and Annexin A5, which anchors La to transiently exposed phosphatidylserine at the surface of fusing osteoclasts. Disappearance of cell-surface La, and the return of full length La to the nuclei of mature, multinucleated osteoclasts, acts as an off switch of their fusion activity. Targeting surface La in a novel explant model of fibrous dysplasia inhibits excessive osteoclast formation characteristic of this disease, highlighting La's potential as a therapeutic target.
    DOI:  https://doi.org/10.1038/s41467-023-36168-x
  38. Nat Commun. 2023 Feb 08. 14(1): 692
    CryoET reveals organelle phenotypes in huntington disease patient iPSC-derived and mouse primary neurons.
    Gong-Her Wu, Charlene Smith-Geater, Jesús G Galaz-Montoya, Yingli Gu, Sanket R Gupte, Ranen Aviner, Patrick G Mitchell, Joy Hsu, Ricardo Miramontes, Keona Q Wang, Nicolette R Geller, Cathy Hou, Cristina Danita, Lydia-Marie Joubert, Michael F Schmid, Serena Yeung, Judith Frydman, William Mobley, Chengbiao Wu, Leslie M Thompson, Wah Chiu.
      Huntington's disease (HD) is caused by an expanded CAG repeat in the huntingtin gene, yielding a Huntingtin protein with an expanded polyglutamine tract. While experiments with patient-derived induced pluripotent stem cells (iPSCs) can help understand disease, defining pathological biomarkers remains challenging. Here, we used cryogenic electron tomography to visualize neurites in HD patient iPSC-derived neurons with varying CAG repeats, and primary cortical neurons from BACHD, deltaN17-BACHD, and wild-type mice. In HD models, we discovered sheet aggregates in double membrane-bound organelles, and mitochondria with distorted cristae and enlarged granules, likely mitochondrial RNA granules. We used artificial intelligence to quantify mitochondrial granules, and proteomics experiments reveal differential protein content in isolated HD mitochondria. Knockdown of Protein Inhibitor of Activated STAT1 ameliorated aberrant phenotypes in iPSC- and BACHD neurons. We show that integrated ultrastructural and proteomic approaches may uncover early HD phenotypes to accelerate diagnostics and the development of targeted therapeutics for HD.
    DOI:  https://doi.org/10.1038/s41467-023-36096-w
  39. Sci Transl Med. 2023 Feb 08. 15(682): eade3782
    Metabolic reprogramming by immune-responsive gene 1 up-regulation improves donor heart preservation and function.
    Ienglam Lei, Wei Huang, Pierre Emmanuel Noly, Suyash Naik, Miriyam Ghali, Liu Liu, Francis D Pagani, Ashraf Abou El Ela, Jordan S Pober, Bertram Pitt, Jeffrey L Platt, Marilia Cascalho, Zhong Wang, Y Eugene Chen, Richard M Mortensen, Paul C Tang.
      Preservation quality of donor hearts is a key determinant of transplant success. Preservation duration beyond 4 hours is associated with primary graft dysfunction (PGD). Given transport time constraints, geographical limitations exist for donor-recipient matching, leading to donor heart underutilization. Here, we showed that metabolic reprogramming through up-regulation of the enzyme immune response gene 1 (IRG1) and its product itaconate improved heart function after prolonged preservation. Irg1 transcript induction was achieved by adding the histone deacetylase (HDAC) inhibitor valproic acid (VPA) to a histidine-tryptophan-ketoglutarate solution used for donor heart preservation. VPA increased acetylated H3K27 occupancy at the IRG1 enhancer and IRG1 transcript expression in human donor hearts. IRG1 converts aconitate to itaconate, which has both anti-inflammatory and antioxidant properties. Accordingly, our studies showed that Irg1 transcript up-regulation by VPA treatment increased nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) in mice, which was accompanied by increased antioxidant protein expression [hemeoxygenase 1 (HO1) and superoxide dismutase 1 (SOD1)]. Deletion of Irg1 in mice (Irg1-/-) negated the antioxidant and cardioprotective effects of VPA. Consistent with itaconate's ability to inhibit succinate dehydrogenase, VPA treatment of human hearts increased itaconate availability and reduced succinate accumulation during preservation. VPA similarly increased IRG1 expression in pig donor hearts and improved its function in an ex vivo cardiac perfusion system both at the clinical 4-hour preservation threshold and at 10 hours. These results suggest that augmentation of cardioprotective immune-metabolomic pathways may be a promising therapeutic strategy for improving donor heart function in transplantation.
    DOI:  https://doi.org/10.1126/scitranslmed.ade3782
  40. Diabetes Care. 2023 Feb 06. pii: dc221833. [Epub ahead of print]
    Polygenic Scores Help Reduce Racial Disparities in Predictive Accuracy of Automated Type 1 Diabetes Classification Algorithms.
    Aaron J Deutsch, Lauren Stalbow, Timothy D Majarian, Josep M Mercader, Alisa K Manning, Jose C Florez, Ruth J F Loos, Miriam S Udler.
       OBJECTIVE: Automated algorithms to identify individuals with type 1 diabetes using electronic health records are increasingly used in biomedical research. It is not known whether the accuracy of these algorithms differs by self-reported race. We investigated whether polygenic scores improve identification of individuals with type 1 diabetes.
    RESEARCH DESIGN AND METHODS: We investigated two large hospital-based biobanks (Mass General Brigham [MGB] and BioMe) and identified individuals with type 1 diabetes using an established automated algorithm. We performed medical record reviews to validate the diagnosis of type 1 diabetes. We implemented two published polygenic scores for type 1 diabetes (developed in individuals of European or African ancestry). We assessed the classification algorithm before and after incorporating polygenic scores.
    RESULTS: The automated algorithm was more likely to incorrectly assign a diagnosis of type 1 diabetes in self-reported non-White individuals than in self-reported White individuals (odds ratio 3.45; 95% CI 1.54-7.69; P = 0.0026). After incorporating polygenic scores into the MGB Biobank, the positive predictive value of the type 1 diabetes algorithm increased from 70 to 97% for self-reported White individuals (meaning that 97% of those predicted to have type 1 diabetes indeed had type 1 diabetes) and from 53 to 100% for self-reported non-White individuals. Similar results were found in BioMe.
    CONCLUSIONS: Automated phenotyping algorithms may exacerbate health disparities because of an increased risk of misclassification of individuals from underrepresented populations. Polygenic scores may be used to improve the performance of phenotyping algorithms and potentially reduce this disparity.
    DOI:  https://doi.org/10.2337/dc22-1833
  41. Cell. 2023 Jan 31. pii: S0092-8674(22)01629-4. [Epub ahead of print]
    Diverse CMT2 neuropathies are linked to aberrant G3BP interactions in stress granules.
    Qinqin Cui, Hongyun Bi, Zhanyun Lv, Qigui Wu, Jianfeng Hua, Bokai Gu, Chanjuan Huo, Mingmin Tang, Yanqin Chen, Chongjiu Chen, Sihan Chen, Xinrui Zhang, Zhangrui Wu, Zhengkai Lao, Nengyin Sheng, Chengyong Shen, Yongdeng Zhang, Zhi-Ying Wu, Zhigang Jin, Peiguo Yang, Huaqing Liu, Jinsong Li, Ge Bai.
      Complex diseases often involve the interplay between genetic and environmental factors. Charcot-Marie-Tooth type 2 neuropathies (CMT2) are a group of genetically heterogeneous disorders, in which similar peripheral neuropathology is inexplicably caused by various mutated genes. Their possible molecular links remain elusive. Here, we found that upon environmental stress, many CMT2-causing mutant proteins adopt similar properties by entering stress granules (SGs), where they aberrantly interact with G3BP and integrate into SG pathways. For example, glycyl-tRNA synthetase (GlyRS) is translocated from the cytoplasm into SGs upon stress, where the mutant GlyRS perturbs the G3BP-centric SG network by aberrantly binding to G3BP. This disrupts SG-mediated stress responses, leading to increased stress vulnerability in motoneurons. Disrupting this aberrant interaction rescues SG abnormalities and alleviates motor deficits in CMT2D mice. These findings reveal a stress-dependent molecular link across diverse CMT2 mutants and provide a conceptual framework for understanding genetic heterogeneity in light of environmental stress.
    Keywords:  Charcot-Marie-Tooth neuropathies; G3BP; axon degeneration; environmental stress; genetic heterogeneity; motor neuron; neuromuscular junction; peripheral neuropathy; phase separation; stress granule
    DOI:  https://doi.org/10.1016/j.cell.2022.12.046
  42. Nat Commun. 2023 Feb 04. 14(1): 615
    Different pathways for engulfment and endocytosis of liquid droplets by nanovesicles.
    Rikhia Ghosh, Vahid Satarifard, Reinhard Lipowsky.
      During endocytosis of nanoparticles by cells, the cellular membranes engulf the particles, thereby forming a closed membrane neck that subsequently undergoes fission. For solid nanoparticles, these endocytic processes have been studied in some detail. Recently, such processes have also been found for liquid and condensate droplets, both in vitro and in vivo. These processes start with the spreading of the droplet onto the membrane followed by partial or complete engulfment of the droplet. Here, we use molecular dynamics simulations to study these processes at the nanoscale, for nano-sized droplets and vesicles. For both partial and complete engulfment, we observe two different endocytic pathways. Complete engulfment leads to a closed membrane neck which may be formed in a circular or strongly non-circular manner. A closed circular neck undergoes fission, thereby generating two nested daughter vesicles whereas a non-circular neck hinders the fission process. Likewise, partial engulfment of larger droplets leads to open membrane necks which can again have a circular or non-circular shape. Two key parameters identified here for these endocytic pathways are the transbilayer stress asymmetry of the vesicle membrane and the positive or negative line tension of the membrane-droplet contact line.
    DOI:  https://doi.org/10.1038/s41467-023-35847-z
  43. J Exp Med. 2023 Apr 03. pii: e20220538. [Epub ahead of print]220(4):
    Remodeling of colon plasma cell repertoire within ulcerative colitis patients.
    Johannes F Scheid, Basak Eraslan, Andrew Hudak, Eric M Brown, Dallis Sergio, Toni M Delorey, Devan Phillips, Ariel Lefkovith, Alison T Jess, Lennard W Duck, Charles O Elson, Hera Vlamakis, Damian R Plichta, Jacques Deguine, Ashwin N Ananthakrishnan, Daniel B Graham, Aviv Regev, Ramnik J Xavier.
      Plasma cells (PCs) constitute a significant fraction of colonic mucosal cells and contribute to inflammatory infiltrates in ulcerative colitis (UC). While gut PCs secrete bacteria-targeting IgA antibodies, their role in UC pathogenesis is unknown. We performed single-cell V(D)J- and RNA-seq on sorted B cells from the colon of healthy individuals and patients with UC. A large fraction of B cell clones is shared between different colon regions, but inflammation in UC broadly disrupts this landscape, causing transcriptomic changes characterized by an increase in the unfolded protein response (UPR) and antigen presentation genes, clonal expansion, and isotype skewing from IgA1 and IgA2 to IgG1. We also directly expressed and assessed the specificity of 152 mAbs from expanded PC clones. These mAbs show low polyreactivity and autoreactivity and instead target both shared bacterial antigens and specific bacterial strains. Altogether, our results characterize the microbiome-specific colon PC response and how its disruption might contribute to inflammation in UC.
    DOI:  https://doi.org/10.1084/jem.20220538
  44. Cell. 2023 Jan 28. pii: S0092-8674(23)00046-6. [Epub ahead of print]
    Structures of LRP2 reveal a molecular machine for endocytosis.
    Andrew Beenken, Gabriele Cerutti, Julia Brasch, Yicheng Guo, Zizhang Sheng, Hediye Erdjument-Bromage, Zainab Aziz, Shelief Y Robbins-Juarez, Estefania Y Chavez, Goran Ahlsen, Phinikoula S Katsamba, Thomas A Neubert, Anthony W P Fitzpatrick, Jonathan Barasch, Lawrence Shapiro.
      The low-density lipoprotein (LDL) receptor-related protein 2 (LRP2 or megalin) is representative of the phylogenetically conserved subfamily of giant LDL receptor-related proteins, which function in endocytosis and are implicated in diseases of the kidney and brain. Here, we report high-resolution cryoelectron microscopy structures of LRP2 isolated from mouse kidney, at extracellular and endosomal pH. The structures reveal LRP2 to be a molecular machine that adopts a conformation for ligand binding at the cell surface and for ligand shedding in the endosome. LRP2 forms a homodimer, the conformational transformation of which is governed by pH-sensitive sites at both homodimer and intra-protomer interfaces. A subset of LRP2 deleterious missense variants in humans appears to impair homodimer assembly. These observations lay the foundation for further understanding the function and mechanism of LDL receptors and implicate homodimerization as a conserved feature of the LRP receptor subfamily.
    Keywords:  CD91; LDL; LRP1; LRP2; cryo-EM; endocytosis; megalin; pH-sensitive; proteinuria; recycling
    DOI:  https://doi.org/10.1016/j.cell.2023.01.016
  45. Cell Metab. 2023 Feb 07. pii: S1550-4131(23)00009-8. [Epub ahead of print]35(2): 299-315.e8
    Sodium perturbs mitochondrial respiration and induces dysfunctional Tregs.
    Beatriz F Côrte-Real, Ibrahim Hamad, Rebeca Arroyo Hornero, Sabrina Geisberger, Joris Roels, Lauren Van Zeebroeck, Aleksandra Dyczko, Marike W van Gisbergen, Henry Kurniawan, Allon Wagner, Nir Yosef, Susanne N Y Weiss, Klaus G Schmetterer, Agnes Schröder, Luka Krampert, Stefanie Haase, Hendrik Bartolomaeus, Niels Hellings, Yvan Saeys, Ludwig J Dubois, Dirk Brenner, Stefan Kempa, David A Hafler, Johannes Stegbauer, Ralf A Linker, Jonathan Jantsch, Dominik N Müller, Markus Kleinewietfeld.
      FOXP3+ regulatory T cells (Tregs) are central for peripheral tolerance, and their deregulation is associated with autoimmunity. Dysfunctional autoimmune Tregs display pro-inflammatory features and altered mitochondrial metabolism, but contributing factors remain elusive. High salt (HS) has been identified to alter immune function and to promote autoimmunity. By investigating longitudinal transcriptional changes of human Tregs, we identified that HS induces metabolic reprogramming, recapitulating features of autoimmune Tregs. Mechanistically, extracellular HS raises intracellular Na+, perturbing mitochondrial respiration by interfering with the electron transport chain (ETC). Metabolic disturbance by a temporary HS encounter or complex III blockade rapidly induces a pro-inflammatory signature and FOXP3 downregulation, leading to long-term dysfunction in vitro and in vivo. The HS-induced effect could be reversed by inhibition of mitochondrial Na+/Ca2+ exchanger (NCLX). Our results indicate that salt could contribute to metabolic reprogramming and that short-term HS encounter perturb metabolic fitness and long-term function of human Tregs with important implications for autoimmunity.
    Keywords:  FOXP3; autoimmunity; high salt; mitochondrial respiration; regulatory T cells
    DOI:  https://doi.org/10.1016/j.cmet.2023.01.009
  46. Science. 2023 Feb 10. 379(6632): 538-539
    Exaggerated responses to a virus long gone.
    Petter Brodin.
      Multisystem inflammatory syndrome in children is caused by abnormal cell activation.
    DOI:  https://doi.org/10.1126/science.adg2776
  47. Nat Cell Biol. 2023 Feb 06.
    Actin cytoskeleton vulnerability to disulfide stress mediates disulfidptosis.
    Xiaoguang Liu, Litong Nie, Yilei Zhang, Yuelong Yan, Chao Wang, Medina Colic, Kellen Olszewski, Amber Horbath, Xiong Chen, Guang Lei, Chao Mao, Shiqi Wu, Li Zhuang, Masha V Poyurovsky, M James You, Traver Hart, Daniel D Billadeau, Junjie Chen, Boyi Gan.
      SLC7A11-mediated cystine uptake suppresses ferroptosis yet promotes cell death under glucose starvation; the nature of the latter cell death remains unknown. Here we show that aberrant accumulation of intracellular disulfides in SLC7A11high cells under glucose starvation induces a previously uncharacterized form of cell death distinct from apoptosis and ferroptosis. We term this cell death disulfidptosis. Chemical proteomics and cell biological analyses showed that glucose starvation in SLC7A11high cells induces aberrant disulfide bonds in actin cytoskeleton proteins and F-actin collapse in a SLC7A11-dependent manner. CRISPR screens and functional studies revealed that inactivation of the WAVE regulatory complex (which promotes actin polymerization and lamellipodia formation) suppresses disulfidptosis, whereas constitutive activation of Rac promotes disulfidptosis. We further show that glucose transporter inhibitors induce disulfidptosis in SLC7A11high cancer cells and suppress SLC7A11high tumour growth. Our results reveal that the susceptibility of the actin cytoskeleton to disulfide stress mediates disulfidptosis and suggest a therapeutic strategy to target disulfidptosis in cancer treatment.
    DOI:  https://doi.org/10.1038/s41556-023-01091-2
  48. Proc Natl Acad Sci U S A. 2023 Feb 14. 120(7): e2212212120
    Cpeb1b-mediated cytoplasmic polyadenylation of shha mRNA modulates zebrafish definitive hematopoiesis.
    Jian Heng, Boyang Shi, Jia-Yi Zhou, Yifan Zhang, Dongyuan Ma, Yun-Gui Yang, Feng Liu.
      During vertebrate embryogenesis, hematopoietic stem and progenitor cell (HSPC) production through endothelial-to-hematopoietic transition requires suitable developmental signals, but how these signals are accurately regulated remains incompletely understood. Cytoplasmic polyadenylation, which is one of the posttranscriptional regulations, plays a crucial role in RNA metabolism. Here, we report that Cpeb1b-mediated cytoplasmic polyadenylation is important for HSPC specification by translational control of Hedgehog (Hh) signaling during zebrafish early development. Cpeb1b is highly expressed in notochord and its deficiency results in defective HSPC production. Mechanistically, Cpeb1b regulates hemogenic endothelium specification by the Hedgehog-Vegf-Notch axis. We demonstrate that the cytoplasmic polyadenylation element motif-dependent interaction between Cpeb1b and shha messenger RNA (mRNA) in the liquid-like condensates, which are induced by Pabpc1b phase separation, is required for cytoplasmic polyadenylation of shha mRNA. Intriguingly, the cytoplasmic polyadenylation regulates translation but not stability of shha mRNA, which further enhances the Shha protein level and Hh signal transduction. Taken together, our findings uncover the role of Cpeb1b-mediated cytoplasmic polyadenylation in HSPC development and provide insights into how posttranscriptional regulation can direct developmental signals with high fidelity to translate them into cell fate transition.
    Keywords:  Cpeb1b; Hedgehog signaling; cytoplasmic polyadenylation; hematopoietic stem and progenitor cell; translational control
    DOI:  https://doi.org/10.1073/pnas.2212212120
  49. Nat Immunol. 2023 Feb 09.
    Mechanisms of myeloid cell entry to the healthy and diseased central nervous system.
    Lukas Amann, Takahiro Masuda, Marco Prinz.
      Myeloid cells in the central nervous system (CNS), such as microglia, CNS-associated macrophages (CAMs), dendritic cells and monocytes, are vital for steady-state immune homeostasis as well as the resolution of tissue damage during brain development or disease-related pathology. The complementary usage of multimodal high-throughput and high-dimensional single-cell technologies along with recent advances in cell-fate mapping has revealed remarkable myeloid cell heterogeneity in the CNS. Despite the establishment of extensive expression profiles revealing myeloid cell multiplicity, the local anatomical conditions for the temporal- and spatial-dependent cellular engraftment are poorly understood. Here we highlight recent discoveries of the context-dependent mechanisms of myeloid cell migration and settlement into distinct subtissular structures in the CNS. These insights offer better understanding of the factors needed for compartment-specific myeloid cell recruitment, integration and residence during development and perturbation, which may lead to better treatment of CNS diseases.
    DOI:  https://doi.org/10.1038/s41590-022-01415-8
  50. Nat Commun. 2023 Feb 04. 14(1): 619
    An engineered variant of MECR reductase reveals indispensability of long-chain acyl-ACPs for mitochondrial respiration.
    M Tanvir Rahman, M Kristian Koski, Joanna Panecka-Hofman, Werner Schmitz, Alexander J Kastaniotis, Rebecca C Wade, Rik K Wierenga, J Kalervo Hiltunen, Kaija J Autio.
      Mitochondrial fatty acid synthesis (mtFAS) is essential for respiratory function. MtFAS generates the octanoic acid precursor for lipoic acid synthesis, but the role of longer fatty acid products has remained unclear. The structurally well-characterized component of mtFAS, human 2E-enoyl-ACP reductase (MECR) rescues respiratory growth and lipoylation defects of a Saccharomyces cerevisiae Δetr1 strain lacking native mtFAS enoyl reductase. To address the role of longer products of mtFAS, we employed in silico molecular simulations to design a MECR variant with a shortened substrate binding cavity. Our in vitro and in vivo analyses indicate that the MECR G165Q variant allows synthesis of octanoyl groups but not long chain fatty acids, confirming the validity of our computational approach to engineer substrate length specificity. Furthermore, our data imply that restoring lipoylation in mtFAS deficient yeast strains is not sufficient to support respiration and that long chain acyl-ACPs generated by mtFAS are required for mitochondrial function.
    DOI:  https://doi.org/10.1038/s41467-023-36358-7
  51. J Hepatol. 2023 Feb 03. pii: S0168-8278(23)00077-6. [Epub ahead of print]
    An adipocentric perspective on the development and progression of non-alcoholic fatty liver disease.
    Eunyoung Lee, Hannelie Korf, Antonio Vidal-Puig.
      White adipose tissue (WAT) is critical in regulating systemic energy homeostasis in combination with the liver. Although each organ has its specialised functions, they must work coordinately through tightly regulated crosstalks to regulate whole-body metabolism. Adipose tissues and the liver are relatively resilient and can adapt to energy surplus by facilitating triglyceride (TG) storage up to a certain threshold level without significant metabolic disturbances. However, lipid storage in WAT beyond a "personalised" adiposity threshold becomes dysfunctional, including metabolic inflexibility, progressive inflammation, and aberrant adipokine secretion. Moreover, the failure of adipose tissue to store and mobilise lipids results in systemic knock-on lipid overload, particularly in the liver. Factors contributing to hepatic lipid overload include lipids released from WAT, dietary fat intake, and enhanced de novo hepatic lipogenesis (DNL). In contrast, extrahepatic mechanisms counteracting toxic hepatic lipid overload entail coordinated compensation by oxidising the surplus of fatty acids in brown adipose tissue and storing fatty acids as triglycerides in white adipose tissue. Failure of these integrated homeostatic mechanisms leads to quantitative increase and qualitative alterations in the lipidome of the liver. Initially, the hepatocytes preferentially accumulate TG species leading to a relatively "benign" non-alcoholic fatty liver (NAFL). However, with time, inflammatory responses ensue, progressing into more severe conditions such as non-alcoholic steatohepatitis (NASH), hepatic cirrhosis, and hepatocellular carcinoma, in some individuals without an early prognostic clue. Here we highlight the pathogenic importance of obesity-induced "adipose tissue failure", resulting in decreased adipose tissue functionality (i.e. fat storage capacity and metabolic flexibility), promoting NAFL/NASH development and progression.
    Keywords:  Adipokines; Adipose tissue biology; Cytokines; Fatty acid flux; Inflammation; Metabolic flexibility; NAFLD
    DOI:  https://doi.org/10.1016/j.jhep.2023.01.024
  52. Science. 2023 Feb 09. eade5750
    Structures of BIRC6-client complexes provide a mechanism of Smac-mediated release of caspases.
    Moritz Hunkeler, Cyrus Y Jin, Eric S Fischer.
      Tight regulation of apoptosis is essential for metazoan development and prevents diseases such as cancer and neurodegeneration. Caspase activation is central to apoptosis and inhibitor of apoptosis (IAP) proteins are the principal actors that restrain caspase activity and are therefore attractive therapeutic targets. IAPs, in turn, are regulated by mitochondria-derived pro-apoptotic factors such as Smac and HtrA2. Through a series of cryo-electron microscopy (cryo-EM) structures of full-length human baculoviral IAP repeat-containing protein 6 (BIRC6) bound to Smac, caspases, and HtrA2, we provide a molecular understanding for BIRC6-mediated caspase inhibition and its release by Smac. The architecture of BIRC6, together with near-irreversible binding of Smac, elucidates how the IAP inhibitor Smac can effectively control a processive ubiquitin ligase to respond to apoptotic stimuli.
    DOI:  https://doi.org/10.1126/science.ade5750
  53. Nature. 2023 Feb 09.
    How a tiny genetic change inflicts old age on young kids.
      
    Keywords:  Medical research
    DOI:  https://doi.org/10.1038/d41586-023-00355-z
  54. Nat Commun. 2023 Feb 10. 14(1): 744
    Intrafusal-fiber LRP4 for muscle spindle formation and maintenance in adult and aged animals.
    Rangjuan Cao, Peng Chen, Hongsheng Wang, Hongyang Jing, Hongsheng Zhang, Guanglin Xing, Bin Luo, Jinxiu Pan, Zheng Yu, Wen-Cheng Xiong, Lin Mei.
      Proprioception is sensed by muscle spindles for precise locomotion and body posture. Unlike the neuromuscular junction (NMJ) for muscle contraction which has been well studied, mechanisms of spindle formation are not well understood. Here we show that sensory nerve terminals are disrupted by the mutation of Lrp4, a gene required for NMJ formation; inducible knockout of Lrp4 in adult mice impairs sensory synapses and movement coordination, suggesting that LRP4 is required for spindle formation and maintenance. LRP4 is critical to the expression of Egr3 during development; in adult mice, it interacts in trans with APP and APLP2 on sensory terminals. Finally, spindle sensory endings and function are impaired in aged mice, deficits that could be diminished by LRP4 expression. These observations uncovered LRP4 as an unexpected regulator of muscle spindle formation and maintenance in adult and aged animals and shed light on potential pathological mechanisms of abnormal muscle proprioception.
    DOI:  https://doi.org/10.1038/s41467-023-36454-8
  55. Nat Commun. 2023 Feb 10. 14(1): 758
    Hedgehog is relayed through dynamic heparan sulfate interactions to shape its gradient.
    Fabian Gude, Jurij Froese, Dominique Manikowski, Daniele Di Iorio, Jean-Noël Grad, Seraphine Wegner, Daniel Hoffmann, Melissa Kennedy, Ralf P Richter, Georg Steffes, Kay Grobe.
      Cellular differentiation is directly determined by concentration gradients of morphogens. As a central model for gradient formation during development, Hedgehog (Hh) morphogens spread away from their source to direct growth and pattern formation in Drosophila wing and eye discs. What is not known is how extracellular Hh spread is achieved and how it translates into precise gradients. Here we show that two separate binding areas located on opposite sides of the Hh molecule can interact directly and simultaneously with two heparan sulfate (HS) chains to temporarily cross-link the chains. Mutated Hh lacking one fully functional binding site still binds HS but shows reduced HS cross-linking. This, in turn, impairs Hhs ability to switch between both chains in vitro and results in striking Hh gradient hypomorphs in vivo. The speed and propensity of direct Hh switching between HS therefore shapes the Hh gradient, revealing a scalable design principle in morphogen-patterned tissues.
    DOI:  https://doi.org/10.1038/s41467-023-36450-y
  56. Cell. 2023 Jan 31. pii: S0092-8674(22)01584-7. [Epub ahead of print]
    Bacteriophages inhibit and evade cGAS-like immune function in bacteria.
    Erin Huiting, Xueli Cao, Jie Ren, Januka S Athukoralage, Zhaorong Luo, Sukrit Silas, Na An, Héloïse Carion, Yu Zhou, James S Fraser, Yue Feng, Joseph Bondy-Denomy.
      A fundamental strategy of eukaryotic antiviral immunity involves the cGAS enzyme, which synthesizes 2',3'-cGAMP and activates the effector STING. Diverse bacteria contain cGAS-like enzymes that produce cyclic oligonucleotides and induce anti-phage activity, known as CBASS. However, this activity has only been demonstrated through heterologous expression. Whether bacteria harboring CBASS antagonize and co-evolve with phages is unknown. Here, we identified an endogenous cGAS-like enzyme in Pseudomonas aeruginosa that generates 3',3'-cGAMP during phage infection, signals to a phospholipase effector, and limits phage replication. In response, phages express an anti-CBASS protein ("Acb2") that forms a hexamer with three 3',3'-cGAMP molecules and reduces phospholipase activity. Acb2 also binds to molecules produced by other bacterial cGAS-like enzymes (3',3'-cUU/UA/UG/AA) and mammalian cGAS (2',3'-cGAMP), suggesting broad inhibition of cGAS-based immunity. Upon Acb2 deletion, CBASS blocks lytic phage replication and lysogenic induction, but rare phages evade CBASS through major capsid gene mutations. Altogether, we demonstrate endogenous CBASS anti-phage function and strategies of CBASS inhibition and evasion.
    Keywords:  CBASS; Pseudomonas aeruginosa; acb; anti-CBASS; anti-phage immunity; bacteriophage; cGAS; capsid; escapers; innate immunity
    DOI:  https://doi.org/10.1016/j.cell.2022.12.041
  57. Nat Commun. 2023 Feb 09. 14(1): 731
    Angiotensin-converting enzyme inhibitor promotes angiogenesis through Sp1/Sp3-mediated inhibition of notch signaling in male mice.
    Hanlin Lu, Peidong Yuan, Xiaoping Ma, Xiuxin Jiang, Shaozhuang Liu, Chang Ma, Sjaak Philipsen, Qunye Zhang, Jianmin Yang, Feng Xu, Cheng Zhang, Yun Zhang, Wencheng Zhang.
      Angiogenesis is a critical pathophysiological process involved in organ growth and various diseases. Transcription factors Sp1/Sp3 are necessary for fetal development and tumor growth. Sp1/Sp3 proteins were downregulated in the capillaries of the gastrocnemius in patients with critical limb ischemia samples. Endothelial-specific Sp1/Sp3 knockout reduces angiogenesis in retinal, pathological, and tumor models and induced activation of the Notch1 pathway. Further, the inactivation of VEGFR2 signaling by Notch1 contributes to the delayed angiogenesis phenotype. Mechanistically, endothelial Sp1 binds to the promoter of Notch1 and inhibits its transcription, which is enhanced by Sp3. The proangiogenic effect of ACEI is abolished in Sp1/Sp3-deletion male mice. We identify USP7 as an ACEI-activated deubiquitinating enzyme that translocated into the nucleus binding to Sp1/Sp3, which are deacetylated by HDAC1. Our findings demonstrate a central role for endothelial USP7-Sp1/Sp3-Notch1 signaling in pathophysiological angiogenesis in response to ACEI treatment.
    DOI:  https://doi.org/10.1038/s41467-023-36409-z
  58. Nat Neurosci. 2023 Feb 09.
    Integrated cardio-behavioral responses to threat define defensive states.
    Jérémy Signoret-Genest, Nina Schukraft, Sara L Reis, Dennis Segebarth, Karl Deisseroth, Philip Tovote.
      Fear and anxiety are brain states that evolved to mediate defensive responses to threats. The defense reaction includes multiple interacting behavioral, autonomic and endocrine adjustments, but their integrative nature is poorly understood. In particular, although threat has been associated with various cardiac changes, there is no clear consensus regarding the relevance of these changes for the integrated defense reaction. Here we identify rapid microstates that are associated with specific behaviors and heart rate dynamics, which are affected by long-lasting macrostates and reflect context-dependent threat levels. In addition, we demonstrate that one of the most commonly used defensive behavioral responses-freezing as measured by immobility-is part of an integrated cardio-behavioral microstate mediated by Chx10+ neurons in the periaqueductal gray. Our framework for systematic integration of cardiac and behavioral readouts presents the basis for a better understanding of complex neural defensive states and their associated systemic functions.
    DOI:  https://doi.org/10.1038/s41593-022-01252-w
  59. Cell Metab. 2023 Feb 07. pii: S1550-4131(23)00008-6. [Epub ahead of print]35(2): 228-230
    Salty Treg cells get out of balance.
    Iosif Papafragkos, Panayotis Verginis.
      Although metabolic rewiring of Treg cells constitutes a hallmark in autoimmune diseases, extrinsic and intrinsic mechanisms that imprint on this re-programming remain poorly understood. In this issue of Cell Metabolism, Côrte-Real and colleagues demonstrate that high salt exposure disturbs the mitochondrial respiration in Treg cells, promoting a pro-inflammatory phenotype, loss of function, and associated breakdown of self-tolerance.
    DOI:  https://doi.org/10.1016/j.cmet.2023.01.008
  60. Nature. 2023 Feb 08.
    The cellular coding of temperature in the mammalian cortex.
    M Vestergaard, M Carta, G Güney, J F A Poulet.
      Temperature is a fundamental sensory modality separate from touch, with dedicated receptor channels and primary afferent neurons for cool and warm1-3. Unlike for other modalities, however, the cortical encoding of temperature remains unknown, with very few cortical neurons reported that respond to non-painful temperature, and the presence of a 'thermal cortex' is debated4-8. Here, using widefield and two-photon calcium imaging in the mouse forepaw system, we identify cortical neurons that respond to cooling and/or warming with distinct spatial and temporal response properties. We observed a representation of cool, but not warm, in the primary somatosensory cortex, but cool and warm in the posterior insular cortex (pIC). The representation of thermal information in pIC is robust and somatotopically arranged, and reversible manipulations show a profound impact on thermal perception. Despite being positioned along the same one-dimensional sensory axis, the encoding of cool and that of warm are distinct, both in highly and broadly tuned neurons. Together, our results show that pIC contains the primary cortical representation of skin temperature and may help explain how the thermal system generates sensations of cool and warm.
    DOI:  https://doi.org/10.1038/s41586-023-05705-5
  61. Nat Commun. 2023 Feb 09. 14(1): 726
    A CpG island-encoded mechanism protects genes from premature transcription termination.
    Amy L Hughes, Aleksander T Szczurek, Jessica R Kelley, Anna Lastuvkova, Anne H Turberfield, Emilia Dimitrova, Neil P Blackledge, Robert J Klose.
      Transcription must be tightly controlled to regulate gene expression and development. However, our understanding of the molecular mechanisms that influence transcription and how these are coordinated in cells to ensure normal gene expression remains rudimentary. Here, by dissecting the function of the SET1 chromatin-modifying complexes that bind to CpG island-associated gene promoters, we discover that they play a specific and essential role in enabling the expression of low to moderately transcribed genes. Counterintuitively, this effect can occur independently of SET1 complex histone-modifying activity and instead relies on an interaction with the RNA Polymerase II-binding protein WDR82. Unexpectedly, we discover that SET1 complexes enable gene expression by antagonising premature transcription termination by the ZC3H4/WDR82 complex at CpG island-associated genes. In contrast, at extragenic sites of transcription, which typically lack CpG islands and SET1 complex occupancy, we show that the activity of ZC3H4/WDR82 is unopposed. Therefore, we reveal a gene regulatory mechanism whereby CpG islands are bound by a protein complex that specifically protects genic transcripts from premature termination, effectively distinguishing genic from extragenic transcription and enabling normal gene expression.
    DOI:  https://doi.org/10.1038/s41467-023-36236-2
  62. J Cell Biol. 2023 Apr 03. pii: e202109090. [Epub ahead of print]222(4):
    Human atlastins are sufficient to drive the fusion of liposomes with a physiological lipid composition.
    Eunhong Jang, Yeojin Moon, So Young Yoon, Joyce Anne R Diaz, Miriam Lee, Naho Ko, Jongseo Park, Soo Hyun Eom, Changwook Lee, Youngsoo Jun.
      The dynamin-like GTPase atlastin is believed to be the minimal machinery required for homotypic endoplasmic reticulum (ER) membrane fusion, mainly because Drosophila atlastin is sufficient to drive liposome fusion. However, it remains unclear whether mammalian atlastins, including the three human atlastins, are sufficient to induce liposome fusion, raising doubts about their major roles in mammalian cells. Here, we show that all human atlastins are sufficient to induce fusion when reconstituted into liposomes with a lipid composition mimicking that of the ER. Although the fusogenic activity of ATL1, which is predominantly expressed in neuronal cells, was weaker than that of ATL2 or ATL3, the addition of M1-spastin, a neuron-specific factor, markedly increased ATL1-mediated liposome fusion. Although we observed efficient fusion between ER microsomes isolated from cultured, non-neuronal cells that predominantly express ATL2-1, an autoinhibited isoform of ATL2, ATL2-1 failed to support liposome fusion by itself as reported previously, indicating that cellular factors enable ATL2-1 to mediate ER fusion in vivo.
    DOI:  https://doi.org/10.1083/jcb.202109090
  63. Nature. 2023 Feb 09.
    How fingerprints get their one-of-a-kind swirls.
    Heidi Ledford.
      
    Keywords:  Biophysics; Developmental biology
    DOI:  https://doi.org/10.1038/d41586-023-00357-x
  64. Nature. 2023 Feb;614(7947): 378-380
    Smart microscopes spot fleeting biology.
    Jyoti Madhusoodanan.
      
    Keywords:  Developmental biology; Imaging; Microscopy; Optics and photonics
    DOI:  https://doi.org/10.1038/d41586-023-00336-2
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