bims-ginsta Biomed News
on Genome instability
Issue of 2024–09–15
forty-five papers selected by
Jinrong Hu, National University of Singapore
  1. Inverse blebs operate as hydraulic pumps during mouse blastocyst formation.
  2. TGF-β and RAS jointly unmask primed enhancers to drive metastasis.
  3. The Polycomb system sustains promoters in a deep OFF state by limiting pre-initiation complex formation to counteract transcription.
  4. Proteome asymmetry in mouse and human embryos before fate specification.
  5. Assembly of a stem cell-derived human postimplantation embryo model.
  6. The cytokinetic midbody mediates asymmetric fate specification at mitotic exit during neural stem cell division.
  7. Quantitative label-free digital holographic imaging of cardiomyocyte optical volume, nucleation, and cell division.
  8. RAPIDASH: Tag-free enrichment of ribosome-associated proteins reveals composition dynamics in embryonic tissue, cancer cells, and macrophages.
  9. Human aneuploid cells depend on the RAF/MEK/ERK pathway for overcoming increased DNA damage.
  10. Spatial organization of translation and translational repression in two phases of germ granules.
  11. Subcellular context-specific tuning of actomyosin ring contractility within a common cytoplasm.
  12. Self-organized tissue mechanics underlie embryonic regulation.
  13. Hyper-recombination in ribosomal DNA is driven by long-range resection-independent RAD51 accumulation.
  14. The central spindle drives anaphase chromosome segregation.
  15. H3K4me2 distinguishes a distinct class of enhancers during the maternal-to-zygotic transition.
  16. Intercellular extrachromosomal DNA copy-number heterogeneity drives neuroblastoma cell state diversity.
  17. Mechanisms of nuclear envelope expansion.
  18. Sustained fertility from first-wave follicle oocytes that pause their growth.
  19. Rejuvenation of aged oocyte through exposure to young follicular microenvironment.
  20. Decoupled transcript and protein concentrations ensure histone homeostasis in different nutrients.
  21. Small-molecule GSDMD agonism in tumors stimulates antitumor immunity without toxicity.
  22. Transcription factor-based transdifferentiation of human embryonic to trophoblast stem cells.
  23. Cross-disorder and disease-specific pathways in dementia revealed by single-cell genomics.
  24. Distinct tumor architectures and microenvironments for the initiation of breast cancer metastasis in the brain.
  25. Tissue-resident memory T cells in epicardial adipose tissue comprise transcriptionally distinct subsets that are modulated in atrial fibrillation.
  26. Gα13 controls pharyngeal endoderm convergence by regulating E-cadherin expression and RhoA activation.
  27. A broadly applicable method for quantifying cardiomyocyte cell division identifies proliferative events following myocardial infarction.
  28. Sex-dependent effects in the aged melanoma tumor microenvironment influence invasion and resistance to targeted therapy.
  29. Comprehensive profiling of migratory primordial germ cells reveals niche-specific differences in non-canonical Wnt and Nodal-Lefty signaling in anterior vs posterior migrants.
  30. TNFSF14+ natural killer cells prevent spontaneous abortion by restricting leucine-mediated decidual stromal cell senescence.
  31. Cage-like complexes that protect folding proteins visualized in cells.
  32. Organoid culture promotes dedifferentiation of mouse myoblasts into stem cells capable of complete muscle regeneration.
  33. Temporally discordant chromatin accessibility and DNA demethylation define short and long-term enhancer regulation during cell fate specification.
  34. Genetic links between ovarian ageing, cancer risk and de novo mutation rates.
  35. The cell rejuvenation atlas: leveraging network biology to identify master regulators of rejuvenation strategies.
  36. An intrinsic mechanism of metabolic tuning promotes cardiac resilience to stress.
  37. Dissecting the regulatory logic of specification and differentiation during vertebrate embryogenesis.
  38. Light-gated integrator for highlighting kinase activity in living cells.
  39. Simultaneous profiling of RNA isoforms and chromatin accessibility of single cells of human retinal organoids.
  40. Custom microfluidic chip design enables cost-effective three-dimensional spatiotemporal transcriptomics with a wide field of view.
  41. Protein kinase N promotes cardiac fibrosis in heart failure by fibroblast-to-myofibroblast conversion.
  42. Modulating the extracellular matrix to treat wound healing defects in Ehlers-Danlos syndrome.
  43. The fitness cost of spurious phosphorylation.
  44. Engineered model of heart tissue repair for exploring fibrotic processes and therapeutic interventions.
  45. Promoter DNA methylation and transcription factor condensation are linked to transcriptional memory in mammalian cells.
  1. Nat Cell Biol. 2024 Sep 11.
    Inverse blebs operate as hydraulic pumps during mouse blastocyst formation.
    Markus F Schliffka, Julien G Dumortier, Diane Pelzer, Arghyadip Mukherjee, Jean-Léon Maître.
      During preimplantation development, mouse embryos form a fluid-filled lumen. Pressurized fluid fractures cell-cell contacts and accumulates into pockets, which coarsen into a single lumen. How the embryo controls intercellular fluid movement during coarsening is unknown. Here we report inverse blebs growing into cells at adhesive contacts. Throughout the embryo we observed hundreds of inverse blebs, each filling with intercellular fluid and retracting within a minute. Inverse blebs grow due to pressure build-up resulting from fluid accumulation and cell-cell adhesion, which locally confines fluid. Inverse blebs retract due to actomyosin contraction, practically pushing fluid within the intercellular space. Importantly, inverse blebs occur infrequently at contacts formed by multiple cells, which effectively serve as fluid sinks. Manipulation of the embryo topology reveals that without sinks inverse blebs pump fluid into one another in futile cycles. We propose that inverse blebs operate as hydraulic pumps to promote luminal coarsening, thereby constituting an instrument used by cells to control fluid movement.
    DOI:  https://doi.org/10.1038/s41556-024-01501-z
  2. Cell. 2024 Aug 29. pii: S0092-8674(24)00905-X. [Epub ahead of print]
    TGF-β and RAS jointly unmask primed enhancers to drive metastasis.
    Jun Ho Lee, Francisco J Sánchez-Rivera, Lan He, Harihar Basnet, Fei Xavier Chen, Elena Spina, Liangji Li, Carles Torner, Jason E Chan, Dig Vijay Kumar Yarlagadda, Jin Suk Park, Carleigh Sussman, Charles M Rudin, Scott W Lowe, Tuomas Tammela, Maria J Macias, Richard P Koche, Joan Massagué.
      Epithelial-to-mesenchymal transitions (EMTs) and extracellular matrix (ECM) remodeling are distinct yet important processes during carcinoma invasion and metastasis. Transforming growth factor β (TGF-β) and RAS, signaling through SMAD and RAS-responsive element-binding protein 1 (RREB1), jointly trigger expression of EMT and fibrogenic factors as two discrete arms of a common transcriptional response in carcinoma cells. Here, we demonstrate that both arms come together to form a program for lung adenocarcinoma metastasis and identify chromatin determinants tying the expression of the constituent genes to TGF-β and RAS inputs. RREB1 localizes to H4K16acK20ac marks in histone H2A.Z-loaded nucleosomes at enhancers in the fibrogenic genes interleukin-11 (IL11), platelet-derived growth factor-B (PDGFB), and hyaluronan synthase 2 (HAS2), as well as the EMT transcription factor SNAI1, priming these enhancers for activation by a SMAD4-INO80 nucleosome remodeling complex in response to TGF-β. These regulatory properties segregate the fibrogenic EMT program from RAS-independent TGF-β gene responses and illuminate the operation and vulnerabilities of a bifunctional program that promotes metastatic outgrowth.
    Keywords:  EMT; RAS; SMAD; TGF-β; chromatin; enhancer; fibrosis; lung adenocarcinoma; metastasis
    DOI:  https://doi.org/10.1016/j.cell.2024.08.014
  3. Nat Cell Biol. 2024 Sep 11.
    The Polycomb system sustains promoters in a deep OFF state by limiting pre-initiation complex formation to counteract transcription.
    Aleksander T Szczurek, Emilia Dimitrova, Jessica R Kelley, Neil P Blackledge, Robert J Klose.
      The Polycomb system has fundamental roles in regulating gene expression during mammalian development. However, how it controls transcription to enable gene repression has remained enigmatic. Here, using rapid degron-based depletion coupled with live-cell transcription imaging and single-particle tracking, we show how the Polycomb system controls transcription in single cells. We discover that the Polycomb system is not a constitutive block to transcription but instead sustains a long-lived deep promoter OFF state, which limits the frequency with which the promoter can enter into a transcribing state. We demonstrate that Polycomb sustains this deep promoter OFF state by counteracting the binding of factors that enable early transcription pre-initiation complex formation and show that this is necessary for gene repression. Together, these important discoveries provide a rationale for how the Polycomb system controls transcription and suggests a universal mechanism that could enable the Polycomb system to constrain transcription across diverse cellular contexts.
    DOI:  https://doi.org/10.1038/s41556-024-01493-w
  4. bioRxiv. 2024 Aug 26. pii: 2024.08.26.609777. [Epub ahead of print]
    Proteome asymmetry in mouse and human embryos before fate specification.
    Lisa K Iwamoto-Stohl, Aleksandra A Petelski, Maciej Meglicki, Audrey Fu, Saad Khan, Harrison Specht, Gray Huffman, Jason Derks, Victoria Jorgensen, Bailey A T Weatherbee, Antonia Weberling, Carlos W Gantner, Rachel S Mandelbaum, Richard J Paulson, Lisa Lam, Ali Ahmady, Estefania Sanchez Vasquez, Nikolai Slavov, Magdalena Zernicka-Goetz.
      Pre-patterning of the embryo, driven by spatially localized factors, is a common feature across several non-mammalian species 1-4 . However, mammals display regulative development and thus it was thought that blastomeres of the embryo do not show such pre-patterning, contributing randomly to the three lineages of the blastocyst: the epiblast, primitive endoderm and trophectoderm that will generate the new organism, the yolk sac and placenta respectively 4-6 . Unexpectedly, early blastomeres of mouse and human embryos have been reported to have distinct developmental fates, potential and heterogeneous abundance of certain transcripts 7-12 . Nevertheless, the extent of the earliest intra-embryo differences remains unclear and controversial. Here, by utilizing multiplexed and label-free single-cell proteomics by mass-spectrometry 13 , we show that 2-cell mouse and human embryos contain an alpha and a beta blastomere as defined by differential abundance of hundreds of proteins exhibiting strong functional enrichment for protein synthesis, transport, and degradation. Such asymmetrically distributed proteins include Gps1 and Nedd8, depletion or overexpression of which in one blastomere of the 2-cell embryo impacts lineage segregation. These protein asymmetries increase at 4-cell stage. Intriguingly, halved mouse zygotes display asymmetric protein abundance that resembles alpha and beta blastomeres, suggesting differential proteome localization already within zygotes. We find that beta blastomeres give rise to a blastocyst with a higher proportion of epiblast cells than alpha blastomeres and that vegetal blastomeres, which are known to have a reduced developmental potential, are more likely to be alpha. Human 2-cell blastomeres also partition into two clusters sharing strong concordance with clusters found in mouse, in terms of differentially abundant proteins and functional enrichment. To our knowledge, this is the first demonstration of intra-zygotic and inter-blastomere proteomic asymmetry in mammals that has a role in lineage segregation.
    DOI:  https://doi.org/10.1101/2024.08.26.609777
  5. Nat Protoc. 2024 Sep 11.
    Assembly of a stem cell-derived human postimplantation embryo model.
    Carlos W Gantner, Bailey A T Weatherbee, Yuntao Wang, Magdalena Zernicka-Goetz.
      The embryonic and extraembryonic tissue interactions underlying human embryogenesis at implantation stages are not currently understood. We have generated a pluripotent stem cell-derived model that mimics aspects of peri-implantation development, allowing tractable experimentation otherwise impossible in the human embryo. Activation of the extraembryonic lineage-specific transcription factors GATA6 and SOX17 (hypoblast factors) or GATA3 and TFAP2C (encoding AP2γ; trophoblast factors) in human embryonic stem (ES) cells drive conversion to extraembryonic-like cells. When combined with wild-type ES cells, self-organized embryo-like structures form in the absence of exogenous factors, termed human inducible embryoids (hiEmbryoids). The epiblast-like domain of hiEmbryoids polarizes and differentiates in response to extraembryonic-secreted extracellular matrix and morphogen cues. Extraembryonic mesenchyme, amnion and primordial germ cells are specified in hiEmbryoids in a stepwise fashion. After establishing stable inducible ES lines and converting ES cells to RSeT culture media, the protocol takes 7-10 d to generate hiEmbryoids. Generation of hiEmbryoids can be performed by researchers with basic expertise in stem cell culture.
    DOI:  https://doi.org/10.1038/s41596-024-01042-7
  6. bioRxiv. 2024 Aug 28. pii: 2024.08.27.609974. [Epub ahead of print]
    The cytokinetic midbody mediates asymmetric fate specification at mitotic exit during neural stem cell division.
    Bryce LaFoya, Rhiannon R Penkert, Kenneth E Prehoda.
      Asymmetric cell division (ACD) is a broadly used mechanism for generating cellular diversity. Molecules known as fate determinants are segregated during ACD to generate distinct sibling cell fates, but determinants should not be activated until fate can be specified asymmetrically. Determinants could be activated after cell division but many animal cells complete division long after mitosis ends, raising the question of how activation could occur at mitotic exit taking advantage of the unique state plasticity at this time point. Here we show that the midbody, a microtubule-rich structure that forms in the intercellular bridge connecting nascent siblings, mediates fate determinant activation at mitotic exit in neural stem cells (NSCs) of the Drosophila larval brain. The fate determinants Prospero (Pros) and Brain tumor (Brat) are sequestered at the NSC membrane at metaphase but are released immediately following nuclear division when the midbody forms, well before cell division completes. The midbody isolates nascent sibling cytoplasms, allowing determinant release from the membrane via the cell cycle phosphatase String, without influencing the fate of the incorrect sibling. Our results identify the midbody as a key facilitator of ACD that allows asymmetric fate determinant activation to be initiated before division.
    DOI:  https://doi.org/10.1101/2024.08.27.609974
  7. J Mol Cell Cardiol. 2024 Sep 07. pii: S0022-2828(24)00145-7. [Epub ahead of print]
    Quantitative label-free digital holographic imaging of cardiomyocyte optical volume, nucleation, and cell division.
    Herman Huang, Sangsoon Park, Ines Ross, Joseph Moreno, Sheamin Khyeam, Jacquelyn Simmons, Guo N Huang, Alexander Y Payumo.
      Cardiac regeneration in newborn rodents depends on the ability of pre-existing cardiomyocytes to proliferate and divide. This capacity is lost within the first week of postnatal development when these cells rapidly switch from hyperplasia to hypertrophy, withdraw from the cell cycle, become binucleated, and increase in size. How these dynamic changes in cell size and nucleation impact cardiomyocyte proliferative potential is not well understood. In this study, we innovate the application of a commercially available digital holographic imaging microscope, the Holomonitor M4, to evaluate the proliferative responses of mononucleated and binucleated cardiomyocytes after CHIR99021 treatment, a model proliferative stimulus. This system enables long-term label-free quantitative tracking of primary cardiomyocyte dynamics in real-time with single-cell resolution. Our results confirm that chemical inhibition of glycogen synthase kinase 3 with CHIR99021 promotes complete cell division of both mononucleated and binucleated cardiomyocytes with high frequency. Quantitative tracking of cardiomyocyte volume dynamics during these proliferative events revealed that both mononucleated and binucleated cardiomyocytes reach a similar size-increase threshold prior to attempted cell division. Binucleated cardiomyocytes attempt to divide with lower frequency than mononucleated cardiomyocytes, which may be associated with inadequate increases in cell size. By defining the interrelationship between cardiomyocyte size, nucleation, and cell cycle control, we may better understand the cellular mechanisms that drive the loss of mammalian cardiac regenerative capacity after birth.
    Keywords:  Cardiomyocyte; Cell culture; Cell division; Digital holographic imaging; Heart; Neonatal rat; Nucleation
    DOI:  https://doi.org/10.1016/j.yjmcc.2024.09.003
  8. Mol Cell. 2024 Sep 05. pii: S1097-2765(24)00700-7. [Epub ahead of print]
    RAPIDASH: Tag-free enrichment of ribosome-associated proteins reveals composition dynamics in embryonic tissue, cancer cells, and macrophages.
    Teodorus Theo Susanto, Victoria Hung, Andrew G Levine, Yuxiang Chen, Craig H Kerr, Yongjin Yoo, Juan A Oses-Prieto, Lisa Fromm, Zijian Zhang, Travis C Lantz, Kotaro Fujii, Marius Wernig, Alma L Burlingame, Davide Ruggero, Maria Barna.
      Ribosomes are emerging as direct regulators of gene expression, with ribosome-associated proteins (RAPs) allowing ribosomes to modulate translation. Nevertheless, a lack of technologies to enrich RAPs across sample types has prevented systematic analysis of RAP identities, dynamics, and functions. We have developed a label-free methodology called RAPIDASH to enrich ribosomes and RAPs from any sample. We applied RAPIDASH to mouse embryonic tissues and identified hundreds of potential RAPs, including Dhx30 and Llph, two forebrain RAPs important for neurodevelopment. We identified a critical role of LLPH in neural development linked to the translation of genes with long coding sequences. In addition, we showed that RAPIDASH can identify ribosome changes in cancer cells. Finally, we characterized ribosome composition remodeling during immune cell activation and observed extensive changes post-stimulation. RAPIDASH has therefore enabled the discovery of RAPs in multiple cell types, tissues, and stimuli and is adaptable to characterize ribosome remodeling in several contexts.
    Keywords:  cancer; embryonic development; macrophages; proteomics; ribosome; ribosome heterogeneity; ribosome-associated proteins; translational control
    DOI:  https://doi.org/10.1016/j.molcel.2024.08.023
  9. Nat Commun. 2024 Sep 09. 15(1): 7772
    Human aneuploid cells depend on the RAF/MEK/ERK pathway for overcoming increased DNA damage.
    Johanna Zerbib, Marica Rosaria Ippolito, Yonatan Eliezer, Giuseppina De Feudis, Eli Reuveni, Anouk Savir Kadmon, Sara Martin, Sonia Viganò, Gil Leor, James Berstler, Julia Muenzner, Michael Mülleder, Emma M Campagnolo, Eldad D Shulman, Tiangen Chang, Carmela Rubolino, Kathrin Laue, Yael Cohen-Sharir, Simone Scorzoni, Silvia Taglietti, Alice Ratti, Chani Stossel, Talia Golan, Francesco Nicassio, Eytan Ruppin, Markus Ralser, Francisca Vazquez, Uri Ben-David, Stefano Santaguida.
      Aneuploidy is a hallmark of human cancer, yet the molecular mechanisms to cope with aneuploidy-induced cellular stresses remain largely unknown. Here, we induce chromosome mis-segregation in non-transformed RPE1-hTERT cells and derive multiple stable clones with various degrees of aneuploidy. We perform a systematic genomic, transcriptomic and proteomic profiling of 6 isogenic clones, using whole-exome DNA, mRNA and miRNA sequencing, as well as proteomics. Concomitantly, we functionally interrogate their cellular vulnerabilities, using genome-wide CRISPR/Cas9 and large-scale drug screens. Aneuploid clones activate the DNA damage response and are more resistant to further DNA damage induction. Aneuploid cells also exhibit elevated RAF/MEK/ERK pathway activity and are more sensitive to clinically-relevant drugs targeting this pathway, and in particular to CRAF inhibition. Importantly, CRAF and MEK inhibition sensitize aneuploid cells to DNA damage-inducing chemotherapies and to PARP inhibitors. We validate these results in human cancer cell lines. Moreover, resistance of cancer patients to olaparib is associated with high levels of RAF/MEK/ERK signaling, specifically in highly-aneuploid tumors. Overall, our study provides a comprehensive resource for genetically-matched karyotypically-stable cells of various aneuploidy states, and reveals a therapeutically-relevant cellular dependency of aneuploid cells.
    DOI:  https://doi.org/10.1038/s41467-024-52176-x
  10. Nat Commun. 2024 Sep 13. 15(1): 8020
    Spatial organization of translation and translational repression in two phases of germ granules.
    Anne Ramat, Ali Haidar, Céline Garret, Martine Simonelig.
      Most RNA-protein condensates are composed of heterogeneous immiscible phases. However, how this multiphase organization contributes to their biological functions remains largely unexplored. Drosophila germ granules, a class of RNA-protein condensates, are the site of mRNA storage and translational activation. Here, using super-resolution microscopy and single-molecule imaging approaches, we show that germ granules have a biphasic organization and that translation occurs in the outer phase and at the surface of the granules. The localization, directionality, and compaction of mRNAs within the granule depend on their translation status, translated mRNAs being enriched in the outer phase with their 5'end oriented towards the surface. Translation is strongly reduced when germ granule biphasic organization is lost. These findings reveal the intimate links between the architecture of RNA-protein condensates and the organization of their different functions, highlighting the functional compartmentalization of these condensates.
    DOI:  https://doi.org/10.1038/s41467-024-52346-x
  11. bioRxiv. 2024 Aug 26. pii: 2024.08.08.607200. [Epub ahead of print]
    Subcellular context-specific tuning of actomyosin ring contractility within a common cytoplasm.
    John B Linehan, Alexandra Zampetaki, Michael E Werner, Bryan Heck, Paul S Maddox, Sebastian Fürthauer, Amy S Maddox.
      The non-muscle actomyosin cytoskeleton generates contractile force through the dynamic rearrangement of its constituent parts. Actomyosin rings are a specialization of the non-muscle actomyosin cytoskeleton that drive cell shape changes during division, wound healing, and other events. Contractile rings throughout phylogeny and in a range of cellular contexts are built from conserved components including non-muscle myosin II (NMMII), actin filaments (F-actin), and crosslinking proteins. However, it is unknown whether diverse actomyosin rings close via a single unifying mechanism. To explore how contractile forces are generated by actomyosin rings, we studied three instances of ring closure within the common cytoplasm of the C. elegans oogenic germline: mitotic cytokinesis of germline stem cells (GSCs), apoptosis of meiotic compartments, and cellularization of oocytes. We found that each ring type closed with unique kinetics, protein density and abundance dynamics. These measurements suggested that the mechanism of contractile force generation varied across the subcellular contexts. Next, we formulated a physical model that related the forces generated by filament-filament interactions to the material properties of these rings that dictate the kinetics of their closure. Using this framework, we related the density of conserved cytoskeletal proteins anillin and NMMII to the kinematics of ring closure. We fitted model rings to in situ measurements to estimate parameters that are currently experimentally inaccessible, such as the asymmetric distribution of protein along the length of F-actin, which occurs naturally due to differences in the dimensions of the crosslinker and NMMII filaments. Our work predicted that the role of NMMII varies across these ring types, due in part to its distribution along F-actin and motoring. Our model also predicted that the degree of contractility and the impact of ring material properties on contractility differs among ring types.
    DOI:  https://doi.org/10.1101/2024.08.08.607200
  12. Nature. 2024 Sep 11.
    Self-organized tissue mechanics underlie embryonic regulation.
    Paolo Caldarelli, Alexander Chamolly, Aurélien Villedieu, Olinda Alegria-Prévot, Carole Phan, Jerome Gros, Francis Corson.
      Early amniote development is highly self-organized, capable of adapting to interference through local and long-range cell-cell interactions. This process, called embryonic regulation1, has been well illustrated in experiments on avian embryos, in which subdividing the epiblast disk into different parts not only redirects cell fates to eventually form a complete and well-proportioned embryo at its original location, but also leads to the self-organization of additional, fully formed embryos2,3 in the other separated parts. The cellular interactions underlying embryonic self-organization are widely believed to be mediated by molecular signals, yet the identity of such signals is unclear. Here, by analysing intact and mechanically perturbed quail embryos, we show that the mechanical forces that drive embryogenesis self-organize, with contractility locally self-activating and the ensuing tension acting as a long-range inhibitor. This mechanical feedback governs the persistent pattern of tissue flows that shape the embryo4-6 and also steers the concomitant emergence of embryonic territories by modulating gene expression, ensuring the formation of a single embryo under normal conditions, yet allowing the emergence of multiple, well-proportioned embryos after perturbations. Thus, mechanical forces act at the core of embryonic self-organization, shaping both tissues and gene expression to robustly yet plastically canalize early development.
    DOI:  https://doi.org/10.1038/s41586-024-07934-8
  13. Nat Commun. 2024 Sep 06. 15(1): 7797
    Hyper-recombination in ribosomal DNA is driven by long-range resection-independent RAD51 accumulation.
    Zita Gál, Stavroula Boukoura, Kezia Catharina Oxe, Sara Badawi, Blanca Nieto, Lea Milling Korsholm, Sille Blangstrup Geisler, Ekaterina Dulina, Anna Vestergaard Rasmussen, Christina Dahl, Wei Lv, Huixin Xu, Xiaoguang Pan, Stefanos Arampatzis, Danai-Eleni Stratou, Panagiotis Galanos, Lin Lin, Per Guldberg, Jiri Bartek, Yonglun Luo, Dorthe H Larsen.
      Ribosomal DNA (rDNA) encodes the ribosomal RNA genes and represents an intrinsically unstable genomic region. However, the underlying mechanisms and implications for genome integrity remain elusive. Here, we use Bloom syndrome (BS), a rare genetic disease characterized by DNA repair defects and hyper-unstable rDNA, as a model to investigate the mechanisms leading to rDNA instability. We find that in Bloom helicase (BLM) proficient cells, the homologous recombination (HR) pathway in rDNA resembles that in nuclear chromatin; it is initiated by resection, replication protein A (RPA) loading and BRCA2-dependent RAD51 filament formation. However, BLM deficiency compromises RPA-loading and BRCA1/2 recruitment to rDNA, but not RAD51 accumulation. RAD51 accumulates at rDNA despite depletion of long-range resection nucleases and rDNA damage results in micronuclei when BLM is absent. In summary, our findings indicate that rDNA is permissive to RAD51 accumulation in the absence of BLM, leading to micronucleation and potentially global genomic instability.
    DOI:  https://doi.org/10.1038/s41467-024-52189-6
  14. bioRxiv. 2024 Aug 30. pii: 2024.08.30.610502. [Epub ahead of print]
    The central spindle drives anaphase chromosome segregation.
    Geng-Yuan Chen, Changfeng Deng, David M Chenoweth, Michael A Lampson.
      Anaphase chromosome segregation depends on forces exerted by spindle microtubules. In the current model, forces on chromosomes are mediated through the spindle poles: sliding of antiparallel microtubules in the central spindle pushes poles apart, while kinetochore microtubule (kMT) depolymerization pulls chromosomes towards the poles. Here we show that the central spindle is directly linked to the chromosomes rather than the poles in anaphase, based on three lines of evidence. Chromosomes in monopolar spindles can move away from the pole, consistent with forces exerted by antiparallel microtubule sliding. In bipolar spindles, kMT depolymerization is constrained by suppressing central spindle sliding, indicating kinetochore linkage to the central spindle. Finally, increasing the rate of kMT depolymerization slows pole separation without increasing chromosome separation velocity. We conclude that central spindle sliding drives anaphase chromosome separation, while kMT depolymerization limits spindle elongation.
    DOI:  https://doi.org/10.1101/2024.08.30.610502
  15. bioRxiv. 2024 Aug 26. pii: 2024.08.26.609713. [Epub ahead of print]
    H3K4me2 distinguishes a distinct class of enhancers during the maternal-to-zygotic transition.
    Matthew D Hurton, Jennifer M Miller, Miler T Lee.
      After egg fertilization, an initially silent embryonic genome is transcriptionally activated during the maternal-to-zygotic transition. In zebrafish, maternal vertebrate pluripotency factors Nanog, Pou5f3 (OCT4 homolog), and Sox19b (SOX2 homolog) (NPS) play essential roles in orchestrating embryonic genome activation, acting as "pioneers" that open condensed chromatin and mediate acquisition of activating histone modifications. However, some embryonic gene transcription still occurs in the absence of these factors, suggesting the existence of other mechanisms regulating genome activation. To identify chromatin signatures of these unknown pathways, we profiled the histone modification landscape of zebrafish embryos using CUT&RUN. Our regulatory map revealed two subclasses of enhancers distinguished by presence or absence of H3K4me2. Enhancers lacking H3K4me2 tend to require NPS factors for de novo activation, while enhancers bearing H3K4me2 are epigenetically bookmarked by DNA hypomethylation to recapitulate gamete activity in the embryo, independent of NPS pioneering. Thus, parallel enhancer activation pathways combine to induce transcriptional reprogramming to pluripotency in the early embryo.
    DOI:  https://doi.org/10.1101/2024.08.26.609713
  16. Cell Rep. 2024 Sep 09. pii: S2211-1247(24)01062-3. [Epub ahead of print]43(9): 114711
    Intercellular extrachromosomal DNA copy-number heterogeneity drives neuroblastoma cell state diversity.
    Maja C Stöber, Rocío Chamorro González, Lotte Brückner, Thomas Conrad, Nadine Wittstruck, Annabell Szymansky, Angelika Eggert, Johannes H Schulte, Richard P Koche, Anton G Henssen, Roland F Schwarz, Kerstin Haase.
      Neuroblastoma exhibits significant inter- and intra-tumor genetic heterogeneity and varying clinical outcomes. Extrachromosomal DNAs (ecDNAs) may drive this heterogeneity by independently segregating during cell division, leading to rapid oncogene amplification. While ecDNA-mediated oncogene amplification is linked to poor prognosis in various cancers, the effects of ecDNA copy-number heterogeneity on intermediate phenotypes are poorly understood. Here, we leverage DNA and RNA sequencing from the same single cells in cell lines and neuroblastoma patients to investigate these effects. By analyzing ecDNA amplicon structures, we reveal extensive intercellular ecDNA copy-number heterogeneity. We also provide direct evidence of how this heterogeneity influences the expression of cargo genes, including MYCN and its downstream targets, and the overall transcriptional state of neuroblastoma cells. Our findings highlight the role of ecDNA copy number in promoting rapid adaptability of cellular states within tumors, underscoring the need for ecDNA-specific treatment strategies to address tumor formation and adaptation.
    Keywords:  CP: Cancer; Extrachromosomal DNA; cell state diversity; copy number; neuroblastoma; single-cell RNA sequencing; tumor heterogeneity
    DOI:  https://doi.org/10.1016/j.celrep.2024.114711
  17. Curr Opin Cell Biol. 2024 Sep 08. pii: S0955-0674(24)00104-2. [Epub ahead of print]91 102425
    Mechanisms of nuclear envelope expansion.
    Christopher Ptak, Saif Rehman, Richard W Wozniak.
      In actively dividing eukaryotic cells, the nuclear envelope membrane (NEM) expands during the cell cycle to accommodate increases in nuclear volume and formation of two nuclei as a cell passes through mitosis to form daughter cells. NEM expansion is driven by glycerophospholipid (GPL) synthesis that is regulated by the lipin family of phosphatidic acid phosphatases (PAPs). How, and when during the cell cycle, PAPs regulate membrane expansion differs between organisms undergoing a closed or open mitosis. Here, we discuss recent studies that shed light on the mechanisms of NE expansion. Moreover, we examine evidence that NEM expansion not only employs GPLs synthesized in the ER but also lipids whose synthesis is regulated by events at the inner nuclear membrane.
    DOI:  https://doi.org/10.1016/j.ceb.2024.102425
  18. bioRxiv. 2024 Aug 28. pii: 2024.08.27.609995. [Epub ahead of print]
    Sustained fertility from first-wave follicle oocytes that pause their growth.
    Bikem Soygur, Eliza A Gaylord, Mariko H Foecke, Steven A Cincotta, Tegan S Horan, Anna Wood, Paula E Cohen, Diana J Laird.
      Ovulation results from the cyclical recruitment of non-renewing, quiescent oocytes for growth. Therefore, the primordial follicles that are established during development from an oocyte encapsulated by granulosa cells are thought to comprise the lifelong ovarian reserve 1-4 . However, using oocyte lineage tracing in mice, we observed that a subset of oocytes recruited for growth in the first juvenile wave remain paused for many months before continuing growth, ovulation, fertilization and development into healthy offspring. This small subset of genetically-labeled fetal oocytes, labeled with Sycp3-CreERT2, is distinguished by earlier entry and slower dynamics of meiotic prophase I. While labeled oocytes were initially found in both primordial follicles and growing follicles of the first wave, they disappeared from primordial follicles by puberty. Unexpectedly, these first-wave labeled growing oocytes persisted throughout reproductive lifespan and contributed to offspring at a steady rate beyond 12 months of age, suggesting that follicles can pause mid-growth for extended periods then successfully resume. These results challenge the conclusion from lineage tracing of granulosa cells that first-wave follicles make a limited contribution to fertility 5 and furthermore suggest that growth-paused oocytes comprise a second and previously unrecognized ovarian reserve.
    DOI:  https://doi.org/10.1101/2024.08.27.609995
  19. Nat Aging. 2024 Sep 09.
    Rejuvenation of aged oocyte through exposure to young follicular microenvironment.
    HaiYang Wang, Zhongwei Huang, Xingyu Shen, Yaelim Lee, XinJie Song, Chang Shu, Lik Hang Wu, Leroy Sivappiragasam Pakkiri, Poh Leong Lim, Xi Zhang, Chester Lee Drum, Jin Zhu, Rong Li.
      Reproductive aging is a major cause of fertility decline, attributed to decreased oocyte quantity and developmental potential. A possible cause is aging of the surrounding follicular somatic cells that support oocyte growth and development by providing nutrients and regulatory factors. Here, by creating chimeric follicles, whereby an oocyte from one follicle was transplanted into and cultured within another follicle whose native oocyte was removed, we show that young oocytes cultured in aged follicles exhibited impeded meiotic maturation and developmental potential, whereas aged oocytes cultured within young follicles were significantly improved in rates of maturation, blastocyst formation and live birth after in vitro fertilization and embryo implantation. This rejuvenation of aged oocytes was associated with enhanced interaction with somatic cells, transcriptomic and metabolomic remodeling, improved mitochondrial function and higher fidelity of meiotic chromosome segregation. These findings provide the basis for a future follicular somatic cell-based therapy to treat female infertility.
    DOI:  https://doi.org/10.1038/s43587-024-00697-x
  20. EMBO J. 2024 Sep 13.
    Decoupled transcript and protein concentrations ensure histone homeostasis in different nutrients.
    Dimitra Chatzitheodoridou, Daniela Bureik, Francesco Padovani, Kalyan V Nadimpalli, Kurt M Schmoller.
      To maintain protein homeostasis in changing nutrient environments, cells must precisely control the amount of their proteins, despite the accompanying changes in cell growth and biosynthetic capacity. As nutrients are major regulators of cell cycle length and progression, a particular challenge arises for the nutrient-dependent regulation of 'cell cycle genes', which are periodically expressed during the cell cycle. One important example are histones, which are needed at a constant histone-to-DNA stoichiometry. Here we show that budding yeast achieves histone homeostasis in different nutrients through a decoupling of transcript and protein abundance. We find that cells downregulate histone transcripts in poor nutrients to avoid toxic histone overexpression, but produce constant amounts of histone proteins through nutrient-specific regulation of translation efficiency. Our findings suggest that this allows cells to balance the need for rapid histone production under fast growth conditions with the tight regulation required to avoid toxic overexpression in poor nutrients.
    Keywords:  Budding Yeast; Cell Cycle; Histones; Nutrients; Protein Homeostasis
    DOI:  https://doi.org/10.1038/s44318-024-00227-w
  21. Cell. 2024 Sep 02. pii: S0092-8674(24)00898-5. [Epub ahead of print]
    Small-molecule GSDMD agonism in tumors stimulates antitumor immunity without toxicity.
    Pietro Fontana, Gang Du, Ying Zhang, Haiwei Zhang, Setu M Vora, Jun Jacob Hu, Ming Shi, Ahmet B Tufan, Liam B Healy, Shiyu Xia, Dian-Jang Lee, Zhouyihan Li, Pilar Baldominos, Heng Ru, Hongbo R Luo, Judith Agudo, Judy Lieberman, Hao Wu.
      Gasdermin-mediated inflammatory cell death (pyroptosis) can activate protective immunity in immunologically cold tumors. Here, we performed a high-throughput screen for compounds that could activate gasdermin D (GSDMD), which is expressed widely in tumors. We identified 6,7-dichloro-2-methylsulfonyl-3-N-tert-butylaminoquinoxaline (DMB) as a direct and selective GSDMD agonist that activates GSDMD pore formation and pyroptosis without cleaving GSDMD. In mouse tumor models, pulsed and low-level pyroptosis induced by DMB suppresses tumor growth without harming GSDMD-expressing immune cells. Protection is immune-mediated and abrogated in mice lacking lymphocytes. Vaccination with DMB-treated cancer cells protects mice from secondary tumor challenge, indicating that immunogenic cell death is induced. DMB treatment synergizes with anti-PD-1. DMB treatment does not alter circulating proinflammatory cytokine or leukocyte numbers or cause weight loss. Thus, our studies reveal a strategy that relies on a low level of tumor cell pyroptosis to induce antitumor immunity and raise the possibility of exploiting pyroptosis without causing overt toxicity.
    Keywords:  GSDMD; GSDMD agonist; antitumor immunity; cancer; checkpoint blockade; gasdermin; immunogenic cell death; immunotherapy; pyroptosis; tumor
    DOI:  https://doi.org/10.1016/j.cell.2024.08.007
  22. Development. 2024 Sep 01. pii: dev202778. [Epub ahead of print]151(17):
    Transcription factor-based transdifferentiation of human embryonic to trophoblast stem cells.
    Paula A Balestrini, Ahmed Abdelbaki, Afshan McCarthy, Liani Devito, Claire E Senner, Alice E Chen, Prabhakaran Munusamy, Paul Blakeley, Kay Elder, Phil Snell, Leila Christie, Paul Serhal, Rabi A Odia, Mahesh Sangrithi, Kathy K Niakan, Norah M E Fogarty.
      During the first week of development, human embryos form a blastocyst composed of an inner cell mass and trophectoderm (TE) cells, the latter of which are progenitors of placental trophoblast. Here, we investigated the expression of transcripts in the human TE from early to late blastocyst stages. We identified enrichment of the transcription factors GATA2, GATA3, TFAP2C and KLF5 and characterised their protein expression dynamics across TE development. By inducible overexpression and mRNA transfection, we determined that these factors, together with MYC, are sufficient to establish induced trophoblast stem cells (iTSCs) from primed human embryonic stem cells. These iTSCs self-renew and recapitulate morphological characteristics, gene expression profiles, and directed differentiation potential, similar to existing human TSCs. Systematic omission of each, or combinations of factors, revealed the crucial importance of GATA2 and GATA3 for iTSC transdifferentiation. Altogether, these findings provide insights into the transcription factor network that may be operational in the human TE and broaden the methods for establishing cellular models of early human placental progenitor cells, which may be useful in the future to model placental-associated diseases.
    Keywords:  Embryonic stem cells; Human embryo; Transdifferentiation; Trophectoderm; Trophoblast stem cells
    DOI:  https://doi.org/10.1242/dev.202778
  23. Cell. 2024 Sep 09. pii: S0092-8674(24)00910-3. [Epub ahead of print]
    Cross-disorder and disease-specific pathways in dementia revealed by single-cell genomics.
    Jessica E Rexach, Yuyan Cheng, Lawrence Chen, Damon Polioudakis, Li-Chun Lin, Vivianne Mitri, Andrew Elkins, Xia Han, Mai Yamakawa, Anna Yin, Daniela Calini, Riki Kawaguchi, Jing Ou, Jerry Huang, Christopher Williams, John Robinson, Stephanie E Gaus, Salvatore Spina, Edward B Lee, Lea T Grinberg, Harry Vinters, John Q Trojanowski, William W Seeley, Dheeraj Malhotra, Daniel H Geschwind.
      The development of successful therapeutics for dementias requires an understanding of their shared and distinct molecular features in the human brain. We performed single-nuclear RNA-seq and ATAC-seq in Alzheimer's disease (AD), frontotemporal dementia (FTD), and progressive supranuclear palsy (PSP), analyzing 41 participants and ∼1 million cells (RNA + ATAC) from three brain regions varying in vulnerability and pathological burden. We identify 32 shared, disease-associated cell types and 14 that are disease specific. Disease-specific cell states represent glial-immune mechanisms and selective neuronal vulnerability impacting layer 5 intratelencephalic neurons in AD, layer 2/3 intratelencephalic neurons in FTD, and layer 5/6 near-projection neurons in PSP. We identify disease-associated gene regulatory networks and cells impacted by causal genetic risk, which differ by disorder. These data illustrate the heterogeneous spectrum of glial and neuronal compositional and gene expression alterations in different dementias and identify therapeutic targets by revealing shared and disease-specific cell states.
    Keywords:  KCNH7; MAFG; NFE2L1; NLGN1; OPCML; PDE1C; drug discovery; functional genomics; multi-omics; tauopathy
    DOI:  https://doi.org/10.1016/j.cell.2024.08.019
  24. Cancer Cell. 2024 Sep 12. pii: S1535-6108(24)00314-3. [Epub ahead of print]
    Distinct tumor architectures and microenvironments for the initiation of breast cancer metastasis in the brain.
    Siting Gan, Danilo G Macalinao, Sayyed Hamed Shahoei, Lin Tian, Xin Jin, Harihar Basnet, Catherine Bibby, James T Muller, Pranita Atri, Evan Seffar, Walid Chatila, Ali Karacay, Pharto Chanda, Anna-Katerina Hadjantonakis, Nikolaus Schultz, Edi Brogi, Tejus A Bale, Nelson S Moss, Rajmohan Murali, Dana Pe'er, Joan Massagué.
      Brain metastasis, a serious complication of cancer, hinges on the initial survival, microenvironment adaptation, and outgrowth of disseminated cancer cells. To understand the early stages of brain colonization, we investigated two prevalent sources of cerebral relapse, triple-negative (TNBC) and HER2+ (HER2BC) breast cancers. Using mouse models and human tissue samples, we found that these tumor types colonize the brain, with a preference for distinctive tumor architectures, stromal interfaces, and autocrine programs. TNBC models tend to form perivascular sheaths with diffusive contact with astrocytes and microglia. In contrast, HER2BC models tend to form compact spheroids driven by autonomous tenascin C production, segregating stromal cells to the periphery. Single-cell transcriptomics of the tumor microenvironment revealed that these architectures evoke differential Alzheimer's disease-associated microglia (DAM) responses and engagement of the GAS6 receptor AXL. The spatial features of the two modes of brain colonization have relevance for leveraging the stroma to treat brain metastasis.
    Keywords:  brain metastasis; breast cancer; extracellular matrix; microglia; tumor architecture; tumor microenvironment
    DOI:  https://doi.org/10.1016/j.ccell.2024.08.015
  25. Nat Cardiovasc Res. 2024 Sep;3(9): 1067-1082
    Tissue-resident memory T cells in epicardial adipose tissue comprise transcriptionally distinct subsets that are modulated in atrial fibrillation.
    Vishal Vyas, Balraj Sandhar, Jack M Keane, Elizabeth G Wood, Hazel Blythe, Aled Jones, Eriomina Shahaj, Silvia Fanti, Jack Williams, Nasrine Metic, Mirjana Efremova, Han Leng Ng, Gayathri Nageswaran, Suzanne Byrne, Niklas Feldhahn, Federica Marelli-Berg, Benny Chain, Andrew Tinker, Malcolm C Finlay, M Paula Longhi.
      Atrial fibrillation (AF) is the most common sustained arrhythmia and carries an increased risk of stroke and heart failure. Here we investigated how the immune infiltrate of human epicardial adipose tissue (EAT), which directly overlies the myocardium, contributes to AF. Flow cytometry analysis revealed an enrichment of tissue-resident memory T (TRM) cells in patients with AF. Cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) and single-cell T cell receptor (TCR) sequencing identified two transcriptionally distinct CD8+ TRM cells that are modulated in AF. Spatial transcriptomic analysis of EAT and atrial tissue identified the border region between the tissues to be a region of intense inflammatory and fibrotic activity, and the addition of TRM populations to atrial cardiomyocytes demonstrated their ability to differentially alter calcium flux as well as activate inflammatory and apoptotic signaling pathways. This study identified EAT as a reservoir of TRM cells that can directly modulate vulnerability to cardiac arrhythmia.
    DOI:  https://doi.org/10.1038/s44161-024-00532-x
  26. Development. 2024 Sep 11. pii: dev.202597. [Epub ahead of print]
    Gα13 controls pharyngeal endoderm convergence by regulating E-cadherin expression and RhoA activation.
    Bo Hu, Joshua Pinzour, Asmi Patel, Faith Rooney, Amie Zerwic, Yuanyuan Gao, Nhan T Nguyen, Huaping Xie, Ding Ye, Fang Lin.
      Pharyngeal endoderm cells undergo convergence and extension (C&E), which is essential for endoderm pouch formation and craniofacial development. Our prior work implicates Gα13/RhoA-mediated signaling in regulating this process, but underlying mechanisms remain unclear. Here, we used endoderm-specific transgenic and Gα13 mutant zebrafish to demonstrate that Gα13 plays a crucial role in pharyngeal endoderm C&E by regulating RhoA activation and E-cadherin expression. We showed that during C&E, endodermal cells gradually establish stable cell-cell contacts, acquire apical-basal polarity, and undergo actomyosin-driven apical constriction, processes that require Gα13. Additionally, we found Gα13-deficient embryos exhibit reduced E-cadherin expression, partially contributing to endoderm C&E defects. Notably, interfering with RhoA function disrupts spatial actomyosin activation without affecting E-cadherin expression. Collectively, our findings identify critical cellular processes for pharyngeal endoderm C&E and reveal that Gα13 controls this through two independent pathways: modulating RhoA activation and regulating E-cadherin expression, thus unveiling intricate mechanisms governing pharyngeal endoderm morphogenesis.
    Keywords:  Convergence and extension; E-cadherin; Gα13; Pharyngeal endoderm; RhoA
    DOI:  https://doi.org/10.1242/dev.202597
  27. Cell Rep Methods. 2024 Aug 30. pii: S2667-2375(24)00238-8. [Epub ahead of print] 100860
    A broadly applicable method for quantifying cardiomyocyte cell division identifies proliferative events following myocardial infarction.
    Samantha K Swift, Alexandra L Purdy, Tyler Buddell, Jerrell J Lovett, Smrithi V Chanjeevaram, Anooj Arkatkar, Caitlin C O'Meara, Michaela Patterson.
      Cardiomyocyte proliferation is a challenging metric to assess. Current methodologies have limitations in detecting the generation of new cardiomyocytes and technical challenges that reduce widespread applicability. Here, we describe an improved cell suspension and imaging-based methodology that can be broadly employed to assess cardiomyocyte cell division in standard laboratories across a multitude of model organisms and experimental conditions. We highlight additional metrics that can be gathered from the same cell preparations to enable additional relevant analyses to be performed. We incorporate additional antibody stains to address potential technical concerns of miscounting. Finally, we employ this methodology with a dual-thymidine analog-labeling approach to a post-infarction murine model, which allowed us to robustly identify unique cycling events, such as cardiomyocytes undergoing multiple rounds of cell division.
    Keywords:  CP: cell biology; cardiomyocyte; cell cycle; endomitosis; heart regeneration; myocardial infarction; proliferation; somatic polyploidy
    DOI:  https://doi.org/10.1016/j.crmeth.2024.100860
  28. Cell. 2024 Sep 04. pii: S0092-8674(24)00904-8. [Epub ahead of print]
    Sex-dependent effects in the aged melanoma tumor microenvironment influence invasion and resistance to targeted therapy.
    Yash Chhabra, Mitchell E Fane, Sneha Pramod, Laura Hüser, Daniel J Zabransky, Vania Wang, Agrani Dixit, Ruzhang Zhao, Edwin Kumah, Megan L Brezka, Kevin Truskowski, Asmita Nandi, Gloria E Marino-Bravante, Alexis E Carey, Naina Gour, Devon A Maranto, Murilo R Rocha, Elizabeth I Harper, Justin Ruiz, Evan J Lipson, Elizabeth M Jaffee, Kristin Bibee, Joel C Sunshine, Hongkai Ji, Ashani T Weeraratna.
      There is documented sex disparity in cutaneous melanoma incidence and mortality, increasing disproportionately with age and in the male sex. However, the underlying mechanisms remain unclear. While biological sex differences and inherent immune response variability have been assessed in tumor cells, the role of the tumor-surrounding microenvironment, contextually in aging, has been overlooked. Here, we show that skin fibroblasts undergo age-mediated, sex-dependent changes in their proliferation, senescence, ROS levels, and stress response. We find that aged male fibroblasts selectively drive an invasive, therapy-resistant phenotype in melanoma cells and promote metastasis in aged male mice by increasing AXL expression. Intrinsic aging in male fibroblasts mediated by EZH2 decline increases BMP2 secretion, which in turn drives the slower-cycling, highly invasive, and therapy-resistant melanoma cell phenotype, characteristic of the aged male TME. Inhibition of BMP2 activity blocks the emergence of invasive phenotypes and sensitizes melanoma cells to BRAF/MEK inhibition.
    Keywords:  DNA damage; aging; epigenetics; fibroblast; melanoma; metastasis; senescence; sex dimorphism; sex disparity; therapy resistance; tumor microenvironment
    DOI:  https://doi.org/10.1016/j.cell.2024.08.013
  29. bioRxiv. 2024 Aug 30. pii: 2024.08.29.610420. [Epub ahead of print]
    Comprehensive profiling of migratory primordial germ cells reveals niche-specific differences in non-canonical Wnt and Nodal-Lefty signaling in anterior vs posterior migrants.
    Rebecca G Jaszczak, Jay W Zussman, Daniel E Wagner, Diana J Laird.
      Mammalian primordial germ cells (PGCs) migrate asynchronously through the embryonic hindgut and dorsal mesentery to reach the gonads. We previously found that interaction with different somatic niches regulates PGC proliferation along the migration route. To characterize transcriptional heterogeneity of migrating PGCs and their niches, we performed single-cell RNA sequencing of 13,262 mouse PGCs and 7,868 surrounding somatic cells during migration (E9.5, E10.5, E11.5) and in anterior versus posterior locations to enrich for leading and lagging migrants. Analysis of PGCs by position revealed dynamic gene expression changes between faster or earlier migrants in the anterior and slower or later migrants in the posterior at E9.5; these differences include migration-associated actin polymerization machinery and epigenetic reprogramming-associated genes. We furthermore identified changes in signaling with various somatic niches, notably strengthened interactions with hindgut epithelium via non-canonical WNT (ncWNT) in posterior PGCs compared to anterior. Reanalysis of a previously published dataset suggests that ncWNT signaling from the hindgut epithelium to early migratory PGCs is conserved in humans. Trajectory inference methods identified putative differentiation trajectories linking cell states across timepoints and from posterior to anterior in our mouse dataset. At E9.5, we mainly observed differences in cell adhesion and actin cytoskeletal dynamics between E9.5 posterior and anterior migrants. At E10.5, we observed divergent gene expression patterns between putative differentiation trajectories from posterior to anterior including Nodal signaling response genes Lefty1, Lefty2, and Pycr2 and reprogramming factors Dnmt1, Prc1, and Tet1. At E10.5, we experimentally validated anterior migrant-specific Lefty1/2 upregulation via whole-mount immunofluorescence staining for LEFTY1/2 proteins, suggesting that elevated autocrine Nodal signaling accompanies the late stages of PGC migration. Together, this positional and temporal atlas of mouse PGCs supports the idea that niche interactions along the migratory route elicit changes in proliferation, actin dynamics, pluripotency, and epigenetic reprogramming.
    DOI:  https://doi.org/10.1101/2024.08.29.610420
  30. EMBO J. 2024 Sep 11.
    TNFSF14+ natural killer cells prevent spontaneous abortion by restricting leucine-mediated decidual stromal cell senescence.
    Jia-Wei Shi, Zhen-Zhen Lai, Wen-Jie Zhou, Hui-Li Yang, Tao Zhang, Jian-Song Sun, Jian-Yuan Zhao, Ming-Qing Li.
      In preparation for a potential pregnancy, the endometrium of the uterus changes into a temporary structure called the decidua. Senescent decidual stromal cells (DSCs) are enriched in the decidua during decidualization, but the underlying mechanisms of this process remain unclear. Here, we performed single-cell RNA transcriptomics on ESCs and DSCs and found that cell senescence during decidualization is accompanied by increased levels of the branched-chain amino acid (BCAA) transporter SLC3A2. Depletion of leucine, one of the branched-chain amino acids, from cultured media decreased senescence, while high leucine diet resulted in increased senescence and high rates of embryo loss in mice. BCAAs induced senescence in DSCs via the p38 MAPK pathway. In contrast, TNFSF14+ decidual natural killer (dNK) cells were found to inhibit DSC senescence by interacting with its ligand TNFRSF14. As in mice fed high-leucine diets, both mice with NK cell depletion and Tnfrsf14-deficient mice with excessive uterine senescence experienced adverse pregnancy outcomes. Further, we found excessive uterine senescence, SLC3A2-mediated BCAA intake, and insufficient TNFRSF14 expression in the decidua of patients with recurrent spontaneous abortion. In summary, this study suggests that dNK cells maintain senescence homeostasis of DSCs via TNFSF14/TNFRSF14, providing a potential therapeutic strategy to prevent DSC senescence-associated spontaneous abortion.
    Keywords:  Abortion; Aging; Decidualization; Leucine; TNFRSF14
    DOI:  https://doi.org/10.1038/s44318-024-00220-3
  31. Nature. 2024 Sep 11.
    Cage-like complexes that protect folding proteins visualized in cells.
      
    Keywords:  Cell biology; Structural biology
    DOI:  https://doi.org/10.1038/d41586-024-02926-0
  32. Nat Biotechnol. 2024 Sep 11.
    Organoid culture promotes dedifferentiation of mouse myoblasts into stem cells capable of complete muscle regeneration.
    Feodor D Price, Mark N Matyas, Andrew R Gehrke, William Chen, Erica A Wolin, Kristina M Holton, Rebecca M Gibbs, Alice Lee, Pooja S Singu, Jeffrey S Sakakeeny, James M Poteracki, Kelsey Goune, Isabella T Pfeiffer, Sarah A Boswell, Peter K Sorger, Mansi Srivastava, Kathleen Lindahl Pfaff, Emanuela Gussoni, Sean M Buchanan, Lee L Rubin.
      Experimental cell therapies for skeletal muscle conditions have shown little success, primarily because they use committed myogenic progenitors rather than true muscle stem cells, known as satellite cells. Here we present a method to generate in vitro-derived satellite cells (idSCs) from skeletal muscle tissue. When transplanted in small numbers into mouse muscle, mouse idSCs fuse into myofibers, repopulate the satellite cell niche, self-renew, support multiple rounds of muscle regeneration and improve force production on par with freshly isolated satellite cells in damaged skeletal muscle. We compared the epigenomic and transcriptional signatures between idSCs, myoblasts and satellite cells and used these signatures to identify core signaling pathways and genes that confer idSC functionality. Finally, from human muscle biopsies, we successfully generated satellite cell-like cells in vitro. After further development, idSCs may provide a scalable source of cells for the treatment of genetic muscle disorders, trauma-induced muscle damage and age-related muscle weakness.
    DOI:  https://doi.org/10.1038/s41587-024-02344-7
  33. bioRxiv. 2024 Aug 27. pii: 2024.08.27.609789. [Epub ahead of print]
    Temporally discordant chromatin accessibility and DNA demethylation define short and long-term enhancer regulation during cell fate specification.
    Lindsey N Guerin, Timothy J Scott, Jacqueline A Yap, Annelie Johansson, Fabio Puddu, Tom Charlesworth, Yilin Yang, Alan J Simmons, Ken S Lau, Rebecca A Ihrie, Emily Hodges.
      Epigenetic mechanisms govern the transcriptional activity of lineage-specifying enhancers; but recent work challenges the dogma that joint chromatin accessibility and DNA demethylation are prerequisites for transcription. To understand this paradox, we established a highly-resolved timeline of DNA demethylation, chromatin accessibility, and transcription factor occupancy during neural progenitor cell differentiation. We show thousands of enhancers undergo rapid, transient accessibility changes associated with distinct periods of transcription factor expression. However, most DNA methylation changes are unidirectional and delayed relative to chromatin dynamics, creating transiently discordant epigenetic states. Genome-wide detection of 5-hydroxymethylcytosine further revealed active demethylation begins ahead of chromatin and transcription factor activity, while enhancer hypomethylation persists long after these activities have dissipated. We demonstrate that these timepoint specific methylation states predict past, present and future chromatin accessibility using machine learning models. Thus, chromatin and DNA methylation collaborate on different timescales to mediate short and long-term enhancer regulation during cell fate specification.
    DOI:  https://doi.org/10.1101/2024.08.27.609789
  34. Nature. 2024 Sep 11.
    Genetic links between ovarian ageing, cancer risk and de novo mutation rates.
    Breast Cancer Association Consortium
      Human genetic studies of common variants have provided substantial insight into the biological mechanisms that govern ovarian ageing1. Here we report analyses of rare protein-coding variants in 106,973 women from the UK Biobank study, implicating genes with effects around five times larger than previously found for common variants (ETAA1, ZNF518A, PNPLA8, PALB2 and SAMHD1). The SAMHD1 association reinforces the link between ovarian ageing and cancer susceptibility1, with damaging germline variants being associated with extended reproductive lifespan and increased all-cause cancer risk in both men and women. Protein-truncating variants in ZNF518A are associated with shorter reproductive lifespan-that is, earlier age at menopause (by 5.61 years) and later age at menarche (by 0.56 years). Finally, using 8,089 sequenced trios from the 100,000 Genomes Project (100kGP), we observe that common genetic variants associated with earlier ovarian ageing associate with an increased rate of maternally derived de novo mutations. Although we were unable to replicate the finding in independent samples from the deCODE study, it is consistent with the expected role of DNA damage response genes in maintaining the genetic integrity of germ cells. This study provides evidence of genetic links between age of menopause and cancer risk.
    DOI:  https://doi.org/10.1038/s41586-024-07931-x
  35. Aging (Albany NY). 2024 Sep 09. null
    The cell rejuvenation atlas: leveraging network biology to identify master regulators of rejuvenation strategies.
    Javier Arcos Hodar, Sascha Jung, Mohamed Soudy, Sybille Barvaux, Antonio Del Sol.
      Current rejuvenation strategies, which range from calorie restriction to in vivo partial reprogramming, only improve a few specific cellular processes. In addition, the molecular mechanisms underlying these approaches are largely unknown, which hinders the design of more holistic cellular rejuvenation strategies. To address this issue, we developed SINGULAR (Single-cell RNA-seq Investigation of Rejuvenation Agents and Longevity), a cell rejuvenation atlas that provides a unified system biology analysis of diverse rejuvenation strategies across multiple organs at single-cell resolution. In particular, we leverage network biology approaches to characterize and compare the effects of each strategy at the level of intracellular signaling, cell-cell communication, and transcriptional regulation. As a result, we identified master regulators orchestrating the rejuvenation response and propose that targeting a combination of them leads to a more holistic improvement of age-dysregulated cellular processes. Thus, the interactive database accompanying SINGULAR is expected to facilitate the future design of synthetic rejuvenation interventions.
    Keywords:  aging; cellular biology; computational biology; database; rejuvenation; scRNA-seq
    DOI:  https://doi.org/10.18632/aging.206105
  36. EMBO Mol Med. 2024 Sep 13.
    An intrinsic mechanism of metabolic tuning promotes cardiac resilience to stress.
    Matteo Sorge, Giulia Savoré, Andrea Gallo, Davide Acquarone, Mauro Sbroggiò, Silvia Velasco, Federica Zamporlini, Saveria Femminò, Enrico Moiso, Giampaolo Morciano, Elisa Balmas, Andrea Raimondi, Gabrielle Nattenberg, Rachele Stefania, Carlo Tacchetti, Angela Maria Rizzo, Paola Corsetto, Alessandra Ghigo, Emilia Turco, Fiorella Altruda, Lorenzo Silengo, Paolo Pinton, Nadia Raffaelli, Nathan J Sniadecki, Claudia Penna, Pasquale Pagliaro, Emilio Hirsch, Chiara Riganti, Guido Tarone, Alessandro Bertero, Mara Brancaccio.
      Defining the molecular mechanisms underlying cardiac resilience is crucial to find effective approaches to protect the heart. A physiologic level of ROS is produced in the heart by fatty acid oxidation, but stressful events can boost ROS and cause mitochondrial dysfunction and cardiac functional impairment. Melusin is a muscle specific chaperone required for myocardial compensatory remodeling during stress. Here we report that Melusin localizes in mitochondria where it binds the mitochondrial trifunctional protein, a key enzyme in fatty acid oxidation, and decreases it activity. Studying both mice and human induced pluripotent stem cell-derived cardiomyocytes, we found that Melusin reduces lipid oxidation in the myocardium and limits ROS generation in steady state and during pressure overload and doxorubicin treatment, preventing mitochondrial dysfunction. Accordingly, the treatment with the lipid oxidation inhibitor Trimetazidine concomitantly with stressful stimuli limits ROS accumulation and prevents long-term heart dysfunction. These findings disclose a physiologic mechanism of metabolic regulation in the heart and demonstrate that a timely restriction of lipid metabolism represents a potential therapeutic strategy to improve cardiac resilience to stress.
    Keywords:  Cardiac Metabolism; Chaperone Proteins; Doxorubicin; Pressure Overload; ROS
    DOI:  https://doi.org/10.1038/s44321-024-00132-z
  37. bioRxiv. 2024 Aug 27. pii: 2024.08.27.609971. [Epub ahead of print]
    Dissecting the regulatory logic of specification and differentiation during vertebrate embryogenesis.
    Jialin Liu, Sebastian M Castillo-Hair, Lucia Y Du, Yiqun Wang, Adam N Carte, Mariona Colomer-Rosell, Christopher Yin, Georg Seelig, Alexander F Schier.
      The interplay between transcription factors and chromatin accessibility regulates cell type diversification during vertebrate embryogenesis. To systematically decipher the gene regulatory logic guiding this process, we generated a single-cell multi-omics atlas of RNA expression and chromatin accessibility during early zebrafish embryogenesis. We developed a deep learning model to predict chromatin accessibility based on DNA sequence and found that a small number of transcription factors underlie cell-type-specific chromatin landscapes. While Nanog is well-established in promoting pluripotency, we discovered a new function in priming the enhancer accessibility of mesendodermal genes. In addition to the classical stepwise mode of differentiation, we describe instant differentiation, where pluripotent cells skip intermediate fate transitions and terminally differentiate. Reconstruction of gene regulatory interactions reveals that this process is driven by a shallow network in which maternally deposited regulators activate a small set of transcription factors that co-regulate hundreds of differentiation genes. Notably, misexpression of these transcription factors in pluripotent cells is sufficient to ectopically activate their targets. This study provides a rich resource for analyzing embryonic gene regulation and reveals the regulatory logic of instant differentiation.
    DOI:  https://doi.org/10.1101/2024.08.27.609971
  38. Nat Commun. 2024 Sep 06. 15(1): 7804
    Light-gated integrator for highlighting kinase activity in living cells.
    Wei Lin, Abhishek Phatarphekar, Yanghao Zhong, Longwei Liu, Hyung-Bae Kwon, William H Gerwick, Yingxiao Wang, Sohum Mehta, Jin Zhang.
      Protein kinases are key signaling nodes that regulate fundamental biological and disease processes. Illuminating kinase signaling from multiple angles can provide deeper insights into disease mechanisms and improve therapeutic targeting. While fluorescent biosensors are powerful tools for visualizing live-cell kinase activity dynamics in real time, new molecular tools are needed that enable recording of transient signaling activities for post hoc analysis and targeted manipulation. Here, we develop a light-gated kinase activity coupled transcriptional integrator (KINACT) that converts dynamic kinase signals into "permanent" fluorescent marks. KINACT enables robust monitoring of kinase activity across scales, accurately recording subcellular PKA activity, highlighting PKA activity distribution in 3D cultures, and identifying PKA activators and inhibitors in high-throughput screens. We further leverage the ability of KINACT to drive signaling effector expression to allow feedback manipulation of the balance of GαsR201C-induced PKA and ERK activation and dissect the mechanisms of oncogenic G protein signaling.
    DOI:  https://doi.org/10.1038/s41467-024-51270-4
  39. Nat Commun. 2024 Sep 13. 15(1): 8022
    Simultaneous profiling of RNA isoforms and chromatin accessibility of single cells of human retinal organoids.
    Shuyao Zhang, Yuhua Xiao, Xinzhi Mo, Xu Chen, Jiawei Zhong, Zheyao Chen, Xu Liu, Yuanhui Qiu, Wangxuan Dai, Jia Chen, Xishan Jin, Guoping Fan, Youjin Hu.
      Single-cell multi-omics sequencing is a powerful approach to analyze complex mechanisms underlying neuronal development and regeneration. However, current methods lack the ability to simultaneously profile RNA alternative splicing and chromatin accessibility at the single-cell level. We develop a technique, single-cell RNA isoform and chromatin accessibility sequencing (scRICA-seq), which demonstrates higher sensitivity and cost-effectiveness compared to existing methods. scRICA-seq can profile both isoforms and chromatin accessibility for up to 10,000 single cells in a single run. Applying this method to human retinal organoids, we construct a multi-omic cell atlas and reveal associations between chromatin accessibility, isoform expression of fate-determining factors, and alternative splicing events in their binding sites. This study provides insights into integrating epigenetics, transcription, and RNA splicing to elucidate the mechanisms underlying retinal neuronal development and fate determination.
    DOI:  https://doi.org/10.1038/s41467-024-52335-0
  40. Nat Genet. 2024 Sep 10.
    Custom microfluidic chip design enables cost-effective three-dimensional spatiotemporal transcriptomics with a wide field of view.
    Junjie Zhu, Kun Pang, Beiyu Hu, Ruiqiao He, Ning Wang, Zewen Jiang, Peifeng Ji, Fangqing Zhao.
      Spatial transcriptomic techniques offer unprecedented insights into the molecular organization of complex tissues. However, integrating cost-effectiveness, high throughput, a wide field of view and compatibility with three-dimensional (3D) volumes has been challenging. Here we introduce microfluidics-assisted grid chips for spatial transcriptome sequencing (MAGIC-seq), a new method that combines carbodiimide chemistry, spatial combinatorial indexing and innovative microfluidics design. This technique allows sensitive and reproducible profiling of diverse tissue types, achieving an eightfold increase in throughput, minimal cost and reduced batch effects. MAGIC-seq breaks conventional microfluidics limits by enhancing barcoding efficiency and enables analysis of whole postnatal mouse sections, providing comprehensive cellular structure elucidation at near single-cell resolution, uncovering transcriptional variations and dynamic trajectories of mouse organogenesis. Our 3D transcriptomic atlas of the developing mouse brain, consisting of 93 sections, reveals the molecular and cellular landscape, serving as a valuable resource for neuroscience and developmental biology. Overall, MAGIC-seq is a high-throughput, cost-effective, large field of view and versatile method for spatial transcriptomic studies.
    DOI:  https://doi.org/10.1038/s41588-024-01906-4
  41. Nat Commun. 2024 Sep 12. 15(1): 7638
    Protein kinase N promotes cardiac fibrosis in heart failure by fibroblast-to-myofibroblast conversion.
    Satoya Yoshida, Tatsuya Yoshida, Kohei Inukai, Katsuhiro Kato, Yoshimitsu Yura, Tomoki Hattori, Atsushi Enomoto, Koji Ohashi, Takahiro Okumura, Noriyuki Ouchi, Haruya Kawase, Nina Wettschureck, Stefan Offermanns, Toyoaki Murohara, Mikito Takefuji.
      Chronic fibrotic tissue disrupts various organ functions. Despite significant advances in therapies, mortality and morbidity due to heart failure remain high, resulting in poor quality of life. Beyond the cardiomyocyte-centric view of heart failure, it is now accepted that alterations in the interstitial extracellular matrix (ECM) also play a major role in the development of heart failure. Here, we show that protein kinase N (PKN) is expressed in cardiac fibroblasts. Furthermore, PKN mediates the conversion of fibroblasts into myofibroblasts, which plays a central role in secreting large amounts of ECM proteins via p38 phosphorylation signaling. Fibroblast-specific deletion of PKN led to a reduction of myocardial fibrotic changes and cardiac dysfunction in mice models of ischemia-reperfusion or heart failure with preserved ejection fraction. Our results indicate that PKN is a therapeutic target for cardiac fibrosis in heart failure.
    DOI:  https://doi.org/10.1038/s41467-024-52068-0
  42. iScience. 2024 Sep 20. 27(9): 110676
    Modulating the extracellular matrix to treat wound healing defects in Ehlers-Danlos syndrome.
    Kindra M Kelly-Scumpia, Maani M Archang, Prabhat K Purbey, Tomohiro Yokota, Rimao Wu, Jackie McCourt, Shen Li, Rachelle H Crosbie, Philip O Scumpia, Arjun Deb.
      Classic Ehlers-Danlos syndrome (cEDS) is a genetic disorder of the connective tissue that is characterized by mutations in genes coding type V collagen. Wound healing defects are characteristic of cEDS and no therapeutic strategies exist. Herein we describe a murine model of cEDS that phenocopies wound healing defects seen in humans. Our model features mice with conditional loss of Col5a1 in Col1a2 + fibroblasts (Col5a1CKO). This model shows that an abnormal extracellular matrix (ECM) characterized by fibrillar disarray, altered mechanical properties, and decreased collagen deposition contribute to the wound healing defect. The cEDS animals exhibit decreased expression of epidermal genes and increased inflammation. Finally, we demonstrate that inhibiting mechanosensitive integrin signaling or by injecting wild-type (WT) fibroblasts into cEDS animals enhances epidermal gene expression, decreases inflammation, and augments wound closure. These findings suggest that cell delivery and/or blocking integrin signaling are potentially therapeutic strategies to rescue wound healing defects in cEDS.
    Keywords:  cell biology; medicine
    DOI:  https://doi.org/10.1016/j.isci.2024.110676
  43. EMBO J. 2024 Sep 10.
    The fitness cost of spurious phosphorylation.
    David Bradley, Alexander Hogrebe, Rohan Dandage, Alexandre K Dubé, Mario Leutert, Ugo Dionne, Alexis Chang, Judit Villén, Christian R Landry.
      The fidelity of signal transduction requires the binding of regulatory molecules to their cognate targets. However, the crowded cell interior risks off-target interactions between proteins that are functionally unrelated. How such off-target interactions impact fitness is not generally known. Here, we use Saccharomyces cerevisiae to inducibly express tyrosine kinases. Because yeast lacks bona fide tyrosine kinases, the resulting tyrosine phosphorylation is biologically spurious. We engineered 44 yeast strains each expressing a tyrosine kinase, and quantitatively analysed their phosphoproteomes. This analysis resulted in ~30,000 phosphosites mapping to ~3500 proteins. The number of spurious pY sites generated correlates strongly with decreased growth, and we predict over 1000 pY events to be deleterious. However, we also find that many of the spurious pY sites have a negligible effect on fitness, possibly because of their low stoichiometry. This result is consistent with our evolutionary analyses demonstrating a lack of phosphotyrosine counter-selection in species with tyrosine kinases. Our results suggest that, alongside the risk for toxicity, the cell can tolerate a large degree of non-functional crosstalk as interaction networks evolve.
    Keywords:  Evolution; Gene Editing; Phosphoproteomics; Protein Kinases; Structural Bioinformatics
    DOI:  https://doi.org/10.1038/s44318-024-00200-7
  44. Nat Commun. 2024 Sep 12. 15(1): 7996
    Engineered model of heart tissue repair for exploring fibrotic processes and therapeutic interventions.
    Pengcheng Yang, Lihang Zhu, Shiya Wang, Jixing Gong, Jonathan Nimal Selvaraj, Lincai Ye, Hanxiao Chen, Yaoyao Zhang, Gongxin Wang, Wanjun Song, Zilong Li, Lin Cai, Hao Zhang, Donghui Zhang.
      Advancements in human-engineered heart tissue have enhanced the understanding of cardiac cellular alteration. Nevertheless, a human model simulating pathological remodeling following myocardial infarction for therapeutic development remains essential. Here we develop an engineered model of myocardial repair that replicates the phased remodeling process, including hypoxic stress, fibrosis, and electrophysiological dysfunction. Transcriptomic analysis identifies nine critical signaling pathways related to cellular fate transitions, leading to the evaluation of seventeen modulators for their therapeutic potential in a mini-repair model. A scoring system quantitatively evaluates the restoration of abnormal electrophysiology, demonstrating that the phased combination of TGFβ inhibitor SB431542, Rho kinase inhibitor Y27632, and WNT activator CHIR99021 yields enhanced functional restoration compared to single factor treatments in both engineered and mouse myocardial infarction model. This engineered heart tissue repair model effectively captures the phased remodeling following myocardial infarction, providing a crucial platform for discovering therapeutic targets for ischemic heart disease.
    DOI:  https://doi.org/10.1038/s41467-024-52221-9
  45. Cell Syst. 2024 Sep 04. pii: S2405-4712(24)00237-0. [Epub ahead of print]
    Promoter DNA methylation and transcription factor condensation are linked to transcriptional memory in mammalian cells.
    Shenqi Fan, Liang Ma, Chengzhi Song, Xu Han, Bijunyao Zhong, Yihan Lin.
      The regulation of genes can be mathematically described by input-output functions that are typically assumed to be time invariant. This fundamental assumption underpins the design of synthetic gene circuits and the quantitative understanding of natural gene regulatory networks. Here, we found that this assumption is challenged in mammalian cells. We observed that a synthetic reporter gene can exhibit unexpected transcriptional memory, leading to a shift in the dose-response curve upon a second induction. Mechanistically, we investigated the cis-dependency of transcriptional memory, revealing the necessity of promoter DNA methylation in establishing memory. Furthermore, we showed that the synthetic transcription factor's effective DNA binding affinity underlies trans-dependency, which is associated with its capacity to undergo biomolecular condensation. These principles enabled modulating memory by perturbing either cis- or trans-regulation of genes. Together, our findings suggest the potential pervasiveness of transcriptional memory and implicate the need to model mammalian gene regulation with time-varying input-output functions. A record of this paper's transparent peer review process is included in the supplemental information.
    Keywords:  CpG methylation; biomolecular condensation; gene regulation; input-output function; massively parallel reporter assay; synthetic biology; transcriptional memory
    DOI:  https://doi.org/10.1016/j.cels.2024.08.007
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