bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2024–11–24
27 papers selected by
Connor Rogerson, University of Cambridge
  1. EMF1 functions as a 3D chromatin modulator in Arabidopsis.
  2. The single-molecule accessibility landscape of newly replicated mammalian chromatin.
  3. KAS-ATAC reveals the genome-wide single-stranded accessible chromatin landscape of the human genome.
  4. A genome-wide screen identifies silencers with distinct chromatin properties and mechanisms of repression.
  5. Phospho-regulation of ASCL1-mediated chromatin opening during cellular reprogramming.
  6. Genome-wide profiling of DNA repair proteins in single cells.
  7. Single-molecule states link transcription factor binding to gene expression.
  8. Crosstalk between paralogs and isoforms influences p63-dependent regulatory element activity.
  9. Cooperative role of LSD1 and CHD7 in regulating differentiation of mouse embryonic stem cells.
  10. Pooled CRISPR screens with joint single-nucleus chromatin accessibility and transcriptome profiling.
  11. Evolving cell states and oncogenic drivers during the progression of IDH-mutant gliomas.
  12. multiDGD: A versatile deep generative model for multi-omics data.
  13. Small-molecule disruption of androgen receptor-dependent chromatin clusters.
  14. Interrogation of the interplay between DNA N6-methyladenosine (6mA) and hypoxia-induced chromatin accessibility by a randomized empirical model (EnrichShuf).
  15. Maternal and zygotic contributions to H3K4me1 chromatin marking during germ layer formation.
  16. Chromatin compaction during confined cell migration induces and reshapes nuclear condensates.
  17. RADIP technology comprehensively identifies H3K27me3-associated RNA-chromatin interactions.
  18. ZBTB7A is a modulator of KDM5-driven transcriptional networks in basal breast cancer.
  19. Interaction of methyl-CpG-binding protein 2 (MeCP2) with distinct enhancers in the mouse cortex.
  20. PTEN depletion reduces H3K27me3 levels to promote epithelial-to-mesenchymal transition in epithelial colorectal cancer cells.
  21. A multi-regional human brain atlas of chromatin accessibility and gene expression facilitates promoter-isoform resolution genetic fine-mapping.
  22. Entangled and non-modular enhancer sequences producing independent spatial activities.
  23. ZBTB24 is a conserved multifaceted transcription factor at genes and centromeres that governs the DNA methylation state and expression of satellite repeats.
  24. PROSER1 modulates DNA demethylation through dual mechanisms to prevent syndromic developmental malformations.
  25. T-follicular helper cells are epigenetically poised to transdifferentiate into T-regulatory type 1 cells.
  26. scCancerExplorer: a comprehensive database for interactively exploring single-cell multi-omics data of human pan-cancer.
  27. Single-cell DNA methylome and 3D genome atlas of the human subcutaneous adipose tissue.
  1. Mol Cell. 2024 Nov 15. pii: S1097-2765(24)00871-2. [Epub ahead of print]
    EMF1 functions as a 3D chromatin modulator in Arabidopsis.
    Jiayue Shu, Linhua Sun, Dingyue Wang, Xiaochang Yin, Minqi Yang, Zhijia Yang, Zheng Gao, Yuehui He, Myriam Calonje, Jinsheng Lai, Xing Wang Deng, Hang He, Yue Zhou.
      It is well known that genome organizers, like mammalian CCCTC-binding factor (CTCF) or Drosophila architectural proteins CP190 and BEAF-32, contribute to the three-dimensional (3D) organization of the genome and ensure normal gene transcription. However, bona fide genome organizers have not been identified in plants. Here, we show that EMBRYONIC FLOWER1 (EMF1) functions as a genome modulator in Arabidopsis. EMF1 interacts with the cohesin component SISTER CHROMATIN COHESION3 (SCC3), and both proteins are enriched at compartment domain (CD) boundaries. Accordingly, emf1 and scc3 show a strength decrease at the CD boundary in which these proteins colocalize. EMF1 maintains CD boundary strength, either independently or in cooperation with histone modifications. Moreover, EMF1 is required to maintain gene-resolution interactions and to block long-range aberrant chromatin loops. These data unveil a key role of EMF1 in regulating 3D chromatin structure.
    Keywords:  3D chromatin structure; EMF1; cohesin; compartment domain; histone modification; modulator
    DOI:  https://doi.org/10.1016/j.molcel.2024.10.031
  2. Cell. 2024 Nov 12. pii: S0092-8674(24)01255-8. [Epub ahead of print]
    The single-molecule accessibility landscape of newly replicated mammalian chromatin.
    Megan S Ostrowski, Marty G Yang, Colin P McNally, Nour J Abdulhay, Simai Wang, Keerthi Renduchintala, Iryna Irkliyenko, Alva Biran, Brandon T L Chew, Ayush D Midha, Emily V Wong, Jonathan Sandoval, Isha H Jain, Anja Groth, Elphège P Nora, Hani Goodarzi, Vijay Ramani.
      We present replication-aware single-molecule accessibility mapping (RASAM), a method to nondestructively measure replication status and protein-DNA interactions on chromatin genome-wide. Using RASAM, we uncover a genome-wide state of single-molecule "hyperaccessibility" post-replication that resolves over several hours. Combining RASAM with cellular models for rapid protein degradation, we demonstrate that histone chaperone CAF-1 reduces nascent chromatin accessibility by filling single-molecular "gaps" and generating closely spaced dinucleosomes on replicated DNA. At cis-regulatory elements, we observe unique modes by which nascent chromatin hyperaccessibility resolves: at CCCTC-binding factor (CTCF)-binding sites, CTCF and nucleosomes compete, reducing CTCF occupancy and motif accessibility post-replication; at active transcription start sites, high chromatin accessibility is maintained, implying rapid re-establishment of nucleosome-free regions. Our study introduces a new paradigm for studying replicated chromatin fiber organization. More broadly, we uncover a unique organization of newly replicated chromatin that must be reset by active processes, providing a substrate for epigenetic reprogramming.
    Keywords:  DNA replication; chromatin; epigenetics; epigenomics; genome architecture; molecular methods; nucleosomes; transcription; transcription factors
    DOI:  https://doi.org/10.1016/j.cell.2024.10.039
  3. Genome Res. 2024 Nov 21. pii: gr.279621.124. [Epub ahead of print]
    KAS-ATAC reveals the genome-wide single-stranded accessible chromatin landscape of the human genome.
    Samuel H Kim, Georgi K Marinov, William Greenleaf.
      Gene regulation in most eukaryotes involves two fundamental physical processes -- alterations in the packaging of the genome by nucleosomes, with active cis-regulatory elements (CREs) generally characterized by an open-chromatin configuration, and the activation of transcription. Mapping these physical properties and biochemical activities genome-wide -- through profiling chromatin accessibility and active transcription -- are key tools used to understand the logic and mechanisms of transcription and its regulation. However, the relationship between these two states has until now not been accessible to simultaneous measurement. To address this, we developed KAS-ATAC, a combination of the KAS-seq (Kethoxal-Assisted SsDNA sequencing and ATAC-seq (Assay for Transposase-Accessible Chromatin using sequencing) methods for mapping single-stranded DNA (and thus active transcription) and chromatin accessibility, respectively, enabling the genome-wide identification of DNA fragments that are simultaneously accessible and contain ssDNA. We use KAS-ATAC to evaluate levels of active transcription over different classes of regulatory elements in the human genome, to estimate the absolute levels of transcribed accessible DNA over CREs, to map the nucleosomal configurations associated with RNA polymerase activities, and to assess transcription factor association with transcribed DNA through transcription factor binding site (TFBS) footprinting. We observe lower levels of transcription over distal enhancers compared to promoters and distinct nucleosomal configurations around transcription initiation sites associated with active transcription. Most TFs associate equally with transcribed and nontranscribed DNA but a few factors specifically do not exhibit footprints over ssDNA-containing fragments. We anticipate KAS-ATAC to continue to derive useful insights into chromatin organization and transcriptional regulation in other contexts in the future.
    DOI:  https://doi.org/10.1101/gr.279621.124
  4. Mol Cell. 2024 Nov 13. pii: S1097-2765(24)00881-5. [Epub ahead of print]
    A genome-wide screen identifies silencers with distinct chromatin properties and mechanisms of repression.
    Lorena Hofbauer, Lisa-Marie Pleyer, Franziska Reiter, Alexander Schleiffer, Anna Vlasova, Leonid Serebreni, Annie Huang, Alexander Stark.
      Differential gene transcription enables development and homeostasis in all animals and is regulated by two major classes of distal cis-regulatory DNA elements (CREs): enhancers and silencers. Although enhancers have been thoroughly characterized, the properties and mechanisms of silencers remain largely unknown. By an unbiased genome-wide functional screen in Drosophila melanogaster S2 cells, we discover a class of silencers that bind one of three transcription factors (TFs) and are generally not included in chromatin-defined CRE catalogs as they mostly lack detectable DNA accessibility. The silencer-binding TF CG11247, which we term Saft, safeguards cell fate decisions in vivo and functions via a highly conserved domain we term zinc-finger-associated C-terminal (ZAC) and the corepressor G9a, independently of G9a's H3K9-methyltransferase activity. Overall, our identification of silencers with unexpected properties and mechanisms has important implications for the understanding and future study of repressive CREs, as well as the functional annotation of animal genomes.
    Keywords:  Drosophila melanogaster; cis-regulatory DNA elements; high-throughput reporter screens; protein domain analysis; silencers; transcription co-factors; transcription factors; transcriptional repression
    DOI:  https://doi.org/10.1016/j.molcel.2024.10.041
  5. Development. 2024 Nov 22. pii: dev.204329. [Epub ahead of print]
    Phospho-regulation of ASCL1-mediated chromatin opening during cellular reprogramming.
    Roberta Azzarelli, Sarah Gillen, Frances Connor, Jethro Lundie-Brown, Francesca Puletti, Rosalind Drummond, Ana Raffaelli, Anna Philpott.
      The proneural transcription factor ASCL1 regulates neurogenesis and drives somatic cell reprogramming into neurons. However, not all cell types can be reprogrammed by ASCL1, raising the questions of what provides competence and how we can overcome barriers to enable directed differentiation. Here, we investigate how levels of ASCL1 and its phosphorylation modulate its activity over progressive lineage restriction of embryonic stem cells. We find that inhibition of ASCL1 phosphorylation enhances reprogramming of both mesodermal and neuroectodermal cells, while pluripotent cells remain refractory to ASCL1-directed neuronal differentiation. By performing RNA-seq and ATAC-seq in neuroectoderm, we find that un(der)phosphorylated ASCL1 causes increased chromatin accessibility at sites proximal to neuronal genes, accompanied by their increased expression. Combined analysis of protein stability and proneural function of phosphomutant and phosphomimetic ASCL1 reveals that protein stability plays only a marginal role in regulating activity, while changes in amino acid charge cannot fully explain enhanced activity of the serine-proline mutant variants of ASCL1. Our work provides new insights into proneural factor activity and regulation and suggests ways to optimize reprogramming protocols in cancer and regenerative medicine.
    Keywords:  Cell fate reprogramming; Chromatin accessibility; Embryonic stem cells; Neurogenesis; Phosphorylation; Proneural transcription factors
    DOI:  https://doi.org/10.1242/dev.204329
  6. Nat Commun. 2024 Nov 21. 15(1): 9918
    Genome-wide profiling of DNA repair proteins in single cells.
    Kim L de Luca, Pim M J Rullens, Magdalena A Karpinska, Sandra S de Vries, Agnieszka Gacek-Matthews, Lőrinc S Pongor, Gaëlle Legube, Joanna W Jachowicz, A Marieke Oudelaar, Jop Kind.
      Accurate repair of DNA damage is critical for maintenance of genomic integrity and cellular viability. Because damage occurs non-uniformly across the genome, single-cell resolution is required for proper interrogation, but sensitive detection has remained challenging. Here, we present a comprehensive analysis of repair protein localization in single human cells using DamID and ChIC sequencing techniques. This study reports genome-wide binding profiles in response to DNA double-strand breaks induced by AsiSI, and explores variability in genomic damage locations and associated repair features in the context of spatial genome organization. By unbiasedly detecting repair factor localization, we find that repair proteins often occupy entire topologically associating domains, mimicking variability in chromatin loop anchoring. Moreover, we demonstrate the formation of multi-way chromatin hubs in response to DNA damage. Notably, larger hubs show increased coordination of repair protein binding, suggesting a preference for cooperative repair mechanisms. Together, our work offers insights into the heterogeneous processes underlying genome stability in single cells.
    DOI:  https://doi.org/10.1038/s41467-024-54159-4
  7. Nature. 2024 Nov 20.
    Single-molecule states link transcription factor binding to gene expression.
    Benjamin R Doughty, Michaela M Hinks, Julia M Schaepe, Georgi K Marinov, Abby R Thurm, Carolina Rios-Martinez, Benjamin E Parks, Yingxuan Tan, Emil Marklund, Danilo Dubocanin, Lacramioara Bintu, William J Greenleaf.
      The binding of multiple transcription factors (TFs) to genomic enhancers drives gene expression in mammalian cells1. However, the molecular details that link enhancer sequence to TF binding, promoter state and transcription levels remain unclear. Here we applied single-molecule footprinting2,3 to measure the simultaneous occupancy of TFs, nucleosomes and other regulatory proteins on engineered enhancer-promoter constructs with variable numbers of TF binding sites for both a synthetic TF and an endogenous TF involved in the type I interferon response. Although TF binding events on nucleosome-free DNA are independent, activation domains recruit cofactors that destabilize nucleosomes, driving observed TF binding cooperativity. Average TF occupancy linearly determines promoter activity, and we decompose TF strength into separable binding and activation terms. Finally, we develop thermodynamic and kinetic models that quantitatively predict both the enhancer binding microstates and gene expression dynamics. This work provides a template for the quantitative dissection of distinct contributors to gene expression, including TF activation domains, concentration, binding affinity, binding site configuration and recruitment of chromatin regulators.
    DOI:  https://doi.org/10.1038/s41586-024-08219-w
  8. Nucleic Acids Res. 2024 Nov 20. pii: gkae1143. [Epub ahead of print]
    Crosstalk between paralogs and isoforms influences p63-dependent regulatory element activity.
    Gabriele Baniulyte, Abby A McCann, Dana L Woodstock, Morgan A Sammons.
      The p53 family of transcription factors (p53, p63 and p73) regulate diverse organismal processes including tumor suppression, maintenance of genome integrity and the development of skin and limbs. Crosstalk between transcription factors with highly similar DNA binding profiles, like those in the p53 family, can dramatically alter gene regulation. While p53 is primarily associated with transcriptional activation, p63 mediates both activation and repression. The specific mechanisms controlling p63-dependent gene regulatory activity are not well understood. Here, we use massively parallel reporter assays (MPRA) to investigate how local DNA sequence context influences p63-dependent transcriptional activity. Most regulatory elements with a p63 response element motif (p63RE) activate transcription, although binding of the p63 paralog, p53, drives a substantial proportion of that activity. p63RE sequence content and co-enrichment with other known activating and repressing transcription factors, including lineage-specific factors, correlates with differential p63RE-mediated activities. p63 isoforms dramatically alter transcriptional behavior, primarily shifting inactive regulatory elements towards high p63-dependent activity. Our analysis provides novel insight into how local sequence and cellular context influences p63-dependent behaviors and highlights the key, yet still understudied, role of transcription factor paralogs and isoforms in controlling gene regulatory element activity.
    DOI:  https://doi.org/10.1093/nar/gkae1143
  9. Sci Rep. 2024 11 18. 14(1): 28495
    Cooperative role of LSD1 and CHD7 in regulating differentiation of mouse embryonic stem cells.
    Sandhya Malla, Carlos Martinez-Gamero, Kanchan Kumari, Cyrinne Achour, Georgios Mermelekas, David Martinez-Delgado, Alba Coego, Diana Guallar, Angel-Carlos Roman, Francesca Aguilo.
      Lysine-specific histone demethylase 1 (LSD1) is a histone demethylase that plays a critical role in epigenetic regulation by removing the methyl group from mono- and di-methylated lysine 4 on histone H3 (H3K4me1/2), acting as a repressor of gene expression. Recently, catalytically independent functions of LSD1, serving as a scaffold for assembling chromatin-regulator and transcription factor complexes, have been identified. Herein, we show for the first time that LSD1 interacts with chromodomain-helicase-DNA-binding protein 7 (CHD7) in mouse embryonic stem cells (ESCs). To further investigate the CHD7-LSD1 crosstalk, we engineered Chd7 and Chd7/Lsd1 knockout (KO) mouse ESCs. We show that CHD7 is dispensable for ESC self-renewal and survival, while Chd7 KO ESCs can differentiate towards embryoid bodies (EBs) with defective expression of ectodermal markers. Intriguingly, Chd7/Lsd1 double KO mouse ESCs exhibit proliferation defects similar to Lsd1 KO ESCs and have lost the capacity to differentiate properly. Furthermore, the increased co-occupancy of H3K4me1 and CHD7 on chromatin following Lsd1 deletion suggests that LSD1 is required for facilitating the proper binding of CHD7 to chromatin and regulating differentiation. Collectively, our results suggest that LSD1 and CHD7 work in concert to modulate gene expression and influence proper cell fate determination.
    DOI:  https://doi.org/10.1038/s41598-024-78920-3
  10. Nat Biotechnol. 2024 Nov 21.
    Pooled CRISPR screens with joint single-nucleus chromatin accessibility and transcriptome profiling.
    Rachel E Yan, Alba Corman, Lyla Katgara, Xiao Wang, Xinhe Xue, Zoran Z Gajic, Richard Sam, Michael Farid, Samuel M Friedman, Jungwook Choo, Ivan Raimondi, Shridar Ganesan, Eugene Katsevich, Jeffrey P Greenfield, Nadia Dahmane, Neville E Sanjana.
      Pooled single-cell CRISPR screens have profiled either gene expression or chromatin accessibility but not both modalities. Here we develop MultiPerturb-seq, a high-throughput CRISPR screening platform with joint single-nucleus chromatin accessibility, transcriptome and guide RNA capture using combinatorial indexing combined with droplet microfluidics to scale throughput and integrate all three modalities. We identify key differentiation genes in a rare pediatric cancer and establish ZNHIT1 as a potential target for cancer reprogramming therapy.
    DOI:  https://doi.org/10.1038/s41587-024-02475-x
  11. Nat Cancer. 2024 Nov 21.
    Evolving cell states and oncogenic drivers during the progression of IDH-mutant gliomas.
    Jingyi Wu, L Nicolas Gonzalez Castro, Sofia Battaglia, Chadi A El Farran, Joshua P D'Antonio, Tyler E Miller, Mario L Suvà, Bradley E Bernstein.
      Isocitrate dehydrogenase (IDH) mutants define a class of gliomas that are initially slow-growing but inevitably progress to fatal disease. To characterize their malignant cell hierarchy, we profiled chromatin accessibility and gene expression across single cells from low-grade and high-grade IDH-mutant gliomas and ascertained their developmental states through a comparison to normal brain cells. We provide evidence that these tumors are initially fueled by slow-cycling oligodendrocyte progenitor cell-like cells. During progression, a more proliferative neural progenitor cell-like population expands, potentially through partial reprogramming of 'permissive' chromatin in progenitors. This transition is accompanied by a switch from methylation-based drivers to genetic ones. In low-grade IDH-mutant tumors or organoids, DNA hypermethylation appears to suppress interferon (IFN) signaling, which is induced by IDH or DNA methyltransferase 1 inhibitors. High-grade tumors frequently lose this hypermethylation and instead acquire genetic alterations that disrupt IFN and other tumor-suppressive programs. Our findings explain how these slow-growing tumors may progress to lethal malignancies and have implications for therapies that target their epigenetic underpinnings.
    DOI:  https://doi.org/10.1038/s43018-024-00865-3
  12. Nat Commun. 2024 Nov 20. 15(1): 10031
    multiDGD: A versatile deep generative model for multi-omics data.
    Viktoria Schuster, Emma Dann, Anders Krogh, Sarah A Teichmann.
      Recent technological advancements in single-cell genomics have enabled joint profiling of gene expression and alternative modalities at unprecedented scale. Consequently, the complexity of multi-omics data sets is increasing massively. Existing models for multi-modal data are typically limited in functionality or scalability, making data integration and downstream analysis cumbersome. We present multiDGD, a scalable deep generative model providing a probabilistic framework to learn shared representations of transcriptome and chromatin accessibility. It shows outstanding performance on data reconstruction without feature selection. We demonstrate on several data sets from human and mouse that multiDGD learns well-clustered joint representations. We further find that probabilistic modeling of sample covariates enables post-hoc data integration without the need for fine-tuning. Additionally, we show that multiDGD can detect statistical associations between genes and regulatory regions conditioned on the learned representations. multiDGD is available as an scverse-compatible package on GitHub.
    DOI:  https://doi.org/10.1038/s41467-024-53340-z
  13. Proc Natl Acad Sci U S A. 2024 Nov 26. 121(48): e2406239121
    Small-molecule disruption of androgen receptor-dependent chromatin clusters.
    Sarah E Kohrt, Emily J Novak, Subhashish Tapadar, Bocheng Wu, Jonathan Strope, Yaw Asante, Hyunmin Kim, Matthew S Chang, Douglas Gurdak, Athar Khalil, Michael Rood, Eric Raftery, Diana Stavreva, Holly M Nguyen, Lisha G Brown, Maddy Ramser, Cody Peer, Warren M Meyers, Nicholas Aboreden, Maharshi Chakravortee, Richard Sallari, Peter S Nelson, Kathleen K Kelly, Thomas G W Graham, Xavier Darzacq, William D Figg, Adegboyega K Oyelere, Eva Corey, Remi Adelaiye-Ogala, Berkley E Gryder.
      Sustained androgen receptor (AR) signaling during relapse is a central driver of metastatic castration-resistant prostate cancer (mCRPC). Current AR antagonists, such as enzalutamide, fail to provide long-term benefit for the mCRPC patients who have dramatic increases in AR expression. Here, we report AR antagonists with efficacy in AR-overexpressing models. These molecules bind to the ligand-binding domain of the AR, promote AR localization to the nucleus, yet potently and selectively down-regulate AR-target genes. The molecules BG-15a and the pharmacokinetically optimized BG-15n elicit a decrease in cell and tumor growth in vitro and in vivo in models of mCRPC. BG-15a/n treatment causes the collapse of chromatin loops between enhancers and promoters at key genes in the AR-driven epigenome. AR binding in the promoter, as well as 3D chromatin clustering, is needed for genes to respond. BG-15a/n represent promising agents for treating patients with relapsed AR-driven mCRPC tumors.
    Keywords:  androgen receptor; chromatin architecture; epigenetics; gene regulation; prostate cancer
    DOI:  https://doi.org/10.1073/pnas.2406239121
  14. Nucleic Acids Res. 2024 Nov 20. pii: gkae1152. [Epub ahead of print]
    Interrogation of the interplay between DNA N6-methyladenosine (6mA) and hypoxia-induced chromatin accessibility by a randomized empirical model (EnrichShuf).
    Joseph Chieh-Yu Lai, Kai-Wen Hsu, Kou-Juey Wu.
      N 6-Methyladenosine (6mA) is an epigenetic mark in eukaryotes regulating development, stress response and tumor progression. METTL4 has been reported as a 6mA methyltransferase induced by hypoxia. The detection and annotation of 6mA signals in mammalian cells have been hampered by the techniques and analytical methods developed so far. Here we developed a 6mA-ChIP-exo-5.1-seq to improve the sensitivity of detecting 6mAs in human cell lines. Furthermore, an EnrichShuf analysis tool for comprehensively comparing 6mA-ChIP-exo-5.1-seq, ATAC-seq, ChIP-seq and RNA-seq has been developed to annotate the functional relevance of 6mA in relation to chromatin accessibility and histone marks. Using a hypoxia-induced 6mA induction system as a model, we showed that hypoxic 6mA signals positively correlated with accessible chromatin regions. These 6mA signals correlate with their regulation by METTL4 under hypoxia, consistent with previous results. 6mAs also co-exist with H3K4me1, a histone mark for enhancers. Further analysis of enhancers using an ABC (active-by-contact) model shows that hypoxia-inducible factor-1α-induced H3K4me3 surrounds the 6mA/H3K4me1 site to augment active enhancers. These results suggest that correlation between 6mA and accessible chromatin regions plays a significant role in enhancer-promoter interactions during hypoxia-induced gene expression.
    DOI:  https://doi.org/10.1093/nar/gkae1152
  15. Dev Biol. 2024 Nov 15. pii: S0012-1606(24)00258-6. [Epub ahead of print]518 8-19
    Maternal and zygotic contributions to H3K4me1 chromatin marking during germ layer formation.
    Kitt D Paraiso, Ira L Blitz, Ken W Y Cho.
      An early step in triploblastic embryo differentiation is the formation of the three germ layers. Maternal pioneer transcription factors (TFs) bind to embryonic enhancers before zygotic genome activation, initiating germ layer specification. While maternal TFs' role in establishing epigenetic marks is known, how early pluripotent cells gain spatially restricted epigenetic identities remains unclear. We show that by the early gastrula stage, H3K4me1-marked regions become distinct in each germ layer, with certain chromatin regions forming high density H3K4me1 marked regions (HDRs). Genes associated with these HDRs are more robustly expressed compared to those associated with low density H3K4me1 marked regions (LDRs) in the genome. This process is driven by the sequential actions of maternal and zygotic factors. Knockdown of key maternal endodermal TFs (Otx1, Vegt and Foxh1) leads to a loss of endodermal H3K4me1 marks in endoderm, with a concurrent emergence of ectodermal and mesodermal marks, indicating a shift in chromatin state. This work highlights the importance of coordinated activities of maternal and zygotic TFs in defining the regionally-resolved and dynamic process of chromatin modification conferred by H3K4me1 in the early Xenopus embryo.
    Keywords:  Canalization; Epigenetics; Gene expression; Histone modification; Robustness; Super enhancers; Transcription factors; Zygotic genome activation
    DOI:  https://doi.org/10.1016/j.ydbio.2024.11.006
  16. Nat Commun. 2024 Nov 18. 15(1): 9964
    Chromatin compaction during confined cell migration induces and reshapes nuclear condensates.
    Jessica Z Zhao, Jing Xia, Clifford P Brangwynne.
      Cell migration through small constrictions during cancer metastasis requires significant deformation of the nucleus, with associated mechanical stress on the nuclear lamina and chromatin. However, how mechanical deformation impacts various subnuclear structures, including protein and nucleic acid-rich biomolecular condensates, is largely unknown. Here, we find that cell migration through confined spaces gives rise to mechanical deformations of the chromatin network, which cause embedded nuclear condensates, including nucleoli and nuclear speckles, to deform and coalesce. Chromatin deformations exhibit differential behavior in the advancing vs. trailing region of the nucleus, with the trailing half being more permissive for de novo condensate formation. We show that this results from increased chromatin heterogeneity, which gives rise to a shift in the binodal phase boundary. Taken together, our findings show how chromatin deformation impacts condensate assembly and properties, which can potentially contribute to cellular mechanosensing.
    DOI:  https://doi.org/10.1038/s41467-024-54120-5
  17. Nucleic Acids Res. 2024 Nov 18. pii: gkae1054. [Epub ahead of print]
    RADIP technology comprehensively identifies H3K27me3-associated RNA-chromatin interactions.
    Xufeng Shu, Masaki Kato, Satoshi Takizawa, Yutaka Suzuki, Piero Carninci.
      Many RNAs associate with chromatin, either directly or indirectly. Several technologies for mapping regions where RNAs interact across the genome have been developed to investigate the function of these RNAs. Obtaining information on the proteins involved in these RNA-chromatin interactions is critical for further analysis. Here, we developed RADIP [RNA and DNA interacting complexes ligated and sequenced (RADICL-seq) with immunoprecipitation], a novel technology that combines RADICL-seq technology with chromatin immunoprecipitation to characterize RNA-chromatin interactions mediated by individual proteins. Building upon the foundational principles of RADICL-seq, RADIP extends its advantages by increasing genomic coverage and unique mapping rate efficiency compared to existing methods. To demonstrate its effectiveness, we applied an anti-H3K27me3 antibody to the RADIP technology and generated libraries from mouse embryonic stem cells (mESCs). We identified a multitude of RNAs, including RNAs from protein-coding genes and non-coding RNAs, that are associated with chromatin via H3K27me3 and that likely facilitate the spread of Polycomb repressive complexes over broad regions of the mammalian genome, thereby affecting gene expression, chromatin structures and pluripotency of mESCs. Our study demonstrates the applicability of RADIP to investigations of the functions of chromatin-associated RNAs.
    DOI:  https://doi.org/10.1093/nar/gkae1054
  18. Cell Rep. 2024 Nov 20. pii: S2211-1247(24)01342-1. [Epub ahead of print]43(12): 114991
    ZBTB7A is a modulator of KDM5-driven transcriptional networks in basal breast cancer.
    Benedetto DiCiaccio, Marco Seehawer, Zheqi Li, Andriana Patmanidis, Triet Bui, Pierre Foidart, Jun Nishida, Clive S D'Santos, Evangelia K Papachristou, Malvina Papanastasiou, Andrew H Reiter, Xintao Qiu, Rong Li, Yijia Jiang, Xiao-Yun Huang, Anton Simeonov, Stephen C Kales, Ganesha Rai, Madhu Lal-Nag, Ajit Jadhav, Myles Brown, Jason S Carroll, Henry W Long, Kornelia Polyak.
      We previously described that the KDM5B histone H3 lysine 4 demethylase is an oncogene in estrogen-receptor-positive breast cancer. Here, we report that KDM5A is amplified and overexpressed in basal breast tumors, and KDM5 inhibition (KDM5i) suppresses the growth of KDM5-amplified breast cancer cell lines. Using CRISPR knockout screens in a basal breast cancer cell line with or without KDM5i, we found that deletion of the ZBTB7A transcription factor and core SAGA complex sensitizes cells to KDM5i, whereas deletion of RHO-GTPases leads to resistance. Chromatin immunoprecipitation sequencing (ChIP-seq) and RNA sequencing (RNA-seq) revealed co-localization of ZBTB7A and KDM5A/B at promoters with high histone H3K4me3 and dependence of KDM5A chromatin binding on ZBTB7A. ZBTB7A knockout altered the transcriptional response to KDM5i at NF-κB targets and mitochondrion-related pathways. High expression of ZBTB7A in triple-negative breast cancer is significantly associated with poor response to neoadjuvant chemotherapy. Our work furthers the understanding of KDM5-mediated gene regulation and identifies mediators of sensitivity to KDM5i.
    Keywords:  CP: Cancer; CRISPR screen; breast cancer; histone demethylase; mitochondrial signaling
    DOI:  https://doi.org/10.1016/j.celrep.2024.114991
  19. Nat Neurosci. 2024 Nov 22.
    Interaction of methyl-CpG-binding protein 2 (MeCP2) with distinct enhancers in the mouse cortex.
    Gyan Prakash Mishra, Eric X Sun, Tiffany Chin, Mandy Eckhardt, Michael E Greenberg, Hume Stroud.
      Mutations in methyl-CpG-binding protein 2 (MeCP2) cause Rett syndrome. MeCP2 is thought to regulate gene transcription by binding to methylated DNA broadly across the genome. Here, using cleavage under target and release under nuclease assays in the adult mouse cortex, we show that MeCP2 strongly binds to specific gene enhancers that we call MeCP2-binding hotspots (MBHs). Unexpectedly, we find that MeCP2 binding to MBHs occurs in a DNA methylation-independent manner at MBHs. Multiple MBH sites surrounding genes mediate the transcriptional repression of genes enriched for neuronal functions. We show that MBHs regulate genes irrespective of genic methylation levels, suggesting that MeCP2 controls transcription via an intragenic methylation-independent mechanism. Hence, disruption of intragenic methylation-independent gene regulation by MeCP2 may in part underlie Rett syndrome.
    DOI:  https://doi.org/10.1038/s41593-024-01808-y
  20. PLoS One. 2024 ;19(11): e0313769
    PTEN depletion reduces H3K27me3 levels to promote epithelial-to-mesenchymal transition in epithelial colorectal cancer cells.
    Ahmed H Ghobashi, Jane W Kimani, Christopher A Ladaika, Heather M O'Hagan.
      Epithelial-to-mesenchymal (EMT) transition is one of the best-known examples of tumor cell plasticity. EMT enhances cancer cell metastasis, which is the main cause of colorectal cancer (CRC)-related mortality. Therefore, understanding underlying molecular mechanisms contributing to the EMT process is crucial to finding druggable targets and more effective therapeutic approaches in CRC. In this study, we demonstrated that phosphatase and tensin homolog (PTEN) knockdown (KD) induces EMT in epithelial CRC, likely through the activation of AKT. PTEN KD modulated chromatin accessibility and reprogrammed gene transcription to mediate EMT in epithelial CRC cells. Active AKT can phosphorylate enhancer of zeste homolog 2 (EZH2) on serine 21, which switches EZH2 from a transcriptional repressor to an activator. Interestingly, PTEN KD reduced the global levels of trimethylation of histone 3 at lysine 27(H3K27me3) in an EZH2-phosphorylation-dependent manner. Additionally, EZH2 phosphorylation at serine 21 reduced the interaction of EZH2 with another polycomb repressive complex 2 (PRC2) component, suppressor of zeste 12 (SUZ12), suggesting that the reduced H3K27me3 levels in PTEN KD cells were due to a disruption of the PRC2 complex. Overall, we demonstrated that PTEN KD modulates changes in gene expression to induce the EMT process in epithelial CRC cells by phosphorylating EZH2 and activates transcription factors such as activator protein 1 (AP1).
    DOI:  https://doi.org/10.1371/journal.pone.0313769
  21. Nat Commun. 2024 Nov 22. 15(1): 10113
    A multi-regional human brain atlas of chromatin accessibility and gene expression facilitates promoter-isoform resolution genetic fine-mapping.
    Pengfei Dong, Liting Song, Jaroslav Bendl, Ruth Misir, Zhiping Shao, Jonathan Edelstien, David A Davis, Vahram Haroutunian, William K Scott, Susanne Acker, Nathan Lawless, Gabriel E Hoffman, John F Fullard, Panos Roussos.
      Brain region- and cell-specific transcriptomic and epigenomic features are associated with heritability for neuropsychiatric traits, but a systematic view, considering cortical and subcortical regions, is lacking. Here, we provide an atlas of chromatin accessibility and gene expression profiles in neuronal and non-neuronal nuclei across 25 distinct human cortical and subcortical brain regions from 6 neurotypical controls. We identified extensive gene expression and chromatin accessibility differences across brain regions, including variation in alternative promoter-isoform usage and enhancer-promoter interactions. Genes with distinct promoter-isoform usage across brain regions were strongly enriched for neuropsychiatric disease risk variants. Moreover, we built enhancer-promoter interactions at promoter-isoform resolution across different brain regions and highlighted the contribution of brain region-specific and promoter-isoform-specific regulation to neuropsychiatric disorders. Including promoter-isoform resolution uncovers additional distal elements implicated in the heritability of diseases, thereby increasing the power to fine-map risk genes. Our results provide a valuable resource for studying molecular regulation across multiple regions of the human brain and underscore the importance of considering isoform information in gene regulation.
    DOI:  https://doi.org/10.1038/s41467-024-54448-y
  22. Sci Adv. 2024 Nov 22. 10(47): eadr9856
    Entangled and non-modular enhancer sequences producing independent spatial activities.
    Mariam Museridze, Stefano Ceolin, Bettina Mühling, Srishti Ramanathan, Olga Barmina, Pallavi Santhi Sekhar, Nicolas Gompel.
      The modularity of transcriptional enhancers is central to our understanding of morphological evolution, allowing specific changes to a gene expression pattern component, without affecting others. Enhancer modularity refers to physically separated stretches of regulatory sequence producing discrete spatiotemporal transcriptional activity. This concept stems from assays that test the sufficiency of a DNA segment to drive spatial reporter expression resembling that of the corresponding gene. Focusing on spatial patterns, it overlooks quantitative aspects of gene expression, underestimating the regulatory sequence actually required to reach full endogenous expression levels. Here, we show that five regulatory activities of the gene yellow in Drosophila, classically described as modular, result from extensively overlapping sequences, with broadly distributed regulatory information. Nevertheless, the independent regulatory activities of these entangled enhancers appear to be nucleated by specific segments that we called enhancer cores. Our work calls for a reappraisal of enhancer definition and properties, as well as of the consequences on regulatory evolution.
    DOI:  https://doi.org/10.1126/sciadv.adr9856
  23. Hum Mol Genet. 2024 Nov 20. pii: ddae163. [Epub ahead of print]
    ZBTB24 is a conserved multifaceted transcription factor at genes and centromeres that governs the DNA methylation state and expression of satellite repeats.
    Giacomo Grillo, Ekaterina Boyarchuk, Seed Mihic, Ivana Ivkovic, Mathilde Bertrand, Alice Jouneau, Thomas Dahlet, Michael Dumas, Michael Weber, Guillaume Velasco, Claire Francastel.
      Since its discovery as a causative gene of the Immunodeficiency with Centromeric instability and Facial anomalies syndrome, ZBTB24 has emerged as a key player in DNA methylation, immunity and development. By extensively analyzing ZBTB24 genomic functions in ICF-relevant mouse and human cellular models, we document here its multiple facets as a transcription factor, with key roles in immune response-related genes expression and also in early embryonic development. Using a constitutive Zbtb24 ICF-like mutant and an auxin-inducible degron system in mouse embryonic stem cells, we showed that ZBTB24 is recruited to centromeric satellite DNA where it is required to establish and maintain the correct DNA methylation patterns through the recruitment of DNMT3B. The ability of ZBTB24 to occupy centromeric satellite DNA is conserved in human cells. Together, our results unveiled an essential and underappreciated role for ZBTB24 at mouse and human centromeric satellite repeat arrays by controlling their DNA methylation and transcription status.
    Keywords:  DNA methylation; ICF syndrome; ZBTB24; centromeric repeats
    DOI:  https://doi.org/10.1093/hmg/ddae163
  24. Genes Dev. 2024 Nov 19.
    PROSER1 modulates DNA demethylation through dual mechanisms to prevent syndromic developmental malformations.
    Anna Fleming, Elena V Knatko, Xiang Li, Ansgar Zoch, Zoe Heckhausen, Stephanie Stransky, Alejandro J Brenes, Simone Sidoli, Petra Hajkova, Dónal O'Carroll, Kasper D Rasmussen.
      The link between DNA methylation and neurodevelopmental disorders is well established. However, how DNA methylation is fine-tuned-ensuring precise gene expression and developmental fidelity-remains poorly understood. PROSER1, a known TET2 interactor, was recently linked to a severe neurodevelopmental disorder. Here, we demonstrate that PROSER1 interacts with all TET enzymes and stabilizes chromatin-bound TET-OGT-PROSER1-DBHS (TOPD) complexes, which regulate DNA demethylation and developmental gene expression. Surprisingly, we found that PROSER1 also sequesters TET enzymes, preventing widespread demethylation and transposable element derepression. Our findings identify PROSER1 as a key factor that both positively and negatively regulates DNA demethylation essential for mammalian neurodevelopment.
    Keywords:  DNA methylation; TET1; TET2; TET3; TOPD; development; neurodevelopmental
    DOI:  https://doi.org/10.1101/gad.352176.124
  25. Elife. 2024 Nov 22. pii: RP97665. [Epub ahead of print]13
    T-follicular helper cells are epigenetically poised to transdifferentiate into T-regulatory type 1 cells.
    Josep Garnica, Patricia Sole, Jun Yamanouchi, Joel Moro, Debajyoti Mondal, Cesar Fandos, Pau Serra, Pere Santamaria.
      Chronic antigenic stimulation can trigger the formation of interleukin 10 (IL-10)-producing T-regulatory type 1 (TR1) cells in vivo. We have recently shown that murine T-follicular helper (TFH) cells are precursors of TR1 cells and that the TFH-to-TR1 cell transdifferentiation process is characterized by the progressive loss and acquisition of opposing transcription factor gene expression programs that evolve through at least one transitional cell stage. Here, we use a broad range of bulk and single-cell transcriptional and epigenetic tools to investigate the epigenetic underpinnings of this process. At the single-cell level, the TFH-to-TR1 cell transition is accompanied by both, downregulation of TFH cell-specific gene expression due to loss of chromatin accessibility, and upregulation of TR1 cell-specific genes linked to chromatin regions that remain accessible throughout the transdifferentiation process, with minimal generation of new open chromatin regions. By interrogating the epigenetic status of accessible TR1 genes on purified TFH and conventional T-cells, we find that most of these genes, including Il10, are already poised for expression at the TFH cell stage. Whereas these genes are closed and hypermethylated in Tconv cells, they are accessible, hypomethylated, and enriched for H3K27ac-marked and hypomethylated active enhancers in TFH cells. These enhancers are enriched for binding sites for the TFH and TR1-associated transcription factors TOX-2, IRF4, and c-MAF. Together, these data suggest that the TR1 gene expression program is genetically imprinted at the TFH cell stage.
    Keywords:  T-follicular helper cells; T-regulatory type 1 cells; TFH cells; TR1 cells; epigenetics; immunology; immunoregulation; inflammation; mouse; pMHCII-based nanomedicines; peptide-major histocompatibility complex class II - based nanomedicines; transdifferentiation
    DOI:  https://doi.org/10.7554/eLife.97665
  26. Nucleic Acids Res. 2024 Nov 18. pii: gkae1100. [Epub ahead of print]
    scCancerExplorer: a comprehensive database for interactively exploring single-cell multi-omics data of human pan-cancer.
    Changzhi Huang, Zekai Liu, Yunlei Guo, Wanchu Wang, Zhen Yuan, Yusheng Guan, Deng Pan, Zhibin Hu, Linhua Sun, Zan Fu, Shuhui Bian.
      Genomic, epigenomic and transcriptomic alterations are hallmarks of cancer cells, and are closely connected. Especially, epigenetic regulation plays a critical role in tumorigenesis and progression. The growing single-cell epigenome data in cancer research provide new opportunities for data mining from a more comprehensive perspective. However, there is still a lack of databases designed for interactively exploring the single-cell multi-omics data of human pan-cancer, especially for the single-cell epigenome data. To fill in the gap, we developed scCancerExplorer, a comprehensive and user-friendly database to facilitate the exploration of the single-cell genome, epigenome (chromatin accessibility and DNA methylation), and transcriptome data of 50 cancer types. Five major modules were provided to explore those data interactively, including 'Integrated multi-omics analysis', 'Single-cell transcriptome', 'Single-cell epigenome', 'Single-cell genome' and 'TCGA analysis'. By simple clicking, users can easily investigate gene expression features, chromatin accessibility patterns, transcription factor activities, DNA methylation states, copy number variations and TCGA survival analysis results. Taken together, scCancerExplorer is distinguished from previous databases with rich and interactive functions for exploring the single-cell multi-omics data of human pan-cancer. It bridges the gap between single-cell multi-omics data and the end-users, and will facilitate progress in the field of cancer research. scCancerExplorer is freely accessible via https://bianlab.cn/scCancerExplorer.
    DOI:  https://doi.org/10.1093/nar/gkae1100
  27. bioRxiv. 2024 Nov 03. pii: 2024.11.02.621694. [Epub ahead of print]
    Single-cell DNA methylome and 3D genome atlas of the human subcutaneous adipose tissue.
    Zeyuan Johnson Chen, Sankha Subhra Das, Asha Kar, Seung Hyuk T Lee, Kevin D Abuhanna, Marcus Alvarez, Mihir G Sukhatme, Kyla Z Gelev, Matthew G Heffel, Yi Zhang, Oren Avram, Elior Rahmani, Sriram Sankararaman, Sini Heinonen, Hilkka Peltoniemi, Eran Halperin, Kirsi H Pietiläinen, Chongyuan Luo, Päivi Pajukanta.
      Human subcutaneous adipose tissue (SAT) contains a diverse array of cell-types; however, the epigenomic landscape among the SAT cell-types has remained elusive. Our integrative analysis of single-cell resolution DNA methylation and chromatin conformation profiles (snm3C-seq), coupled with matching RNA expression (snRNA-seq), systematically cataloged the epigenomic, 3D topology, and transcriptomic dynamics across the SAT cell-types. We discovered that the SAT CG methylation (mCG) landscape is characterized by pronounced hyper-methylation in myeloid cells and hypo-methylation in adipocytes and adipose stem and progenitor cells (ASPCs), driving nearly half of the 705,063 detected differentially methylated regions (DMRs). In addition to the enriched cell-type-specific transcription factor binding motifs, we identified TET1 and DNMT3A as plausible candidates for regulating cell-type level mCG profiles. Furthermore, we observed that global mCG profiles closely correspond to SAT lineage, which is also reflected in cell-type-specific chromosome compartmentalization. Adipocytes, in particular, display significantly more short-range chromosomal interactions, facilitating the formation of complex local 3D genomic structures that regulate downstream transcriptomic activity, including those associated with adipogenesis. Finally, we discovered that variants in cell-type level DMRs and A compartments significantly predict and are enriched for variance explained in abdominal obesity. Together, our multimodal study characterizes human SAT epigenomic landscape at the cell-type resolution and links partitioned polygenic risk of abdominal obesity to SAT epigenome.
    DOI:  https://doi.org/10.1101/2024.11.02.621694
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