bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2020‒11‒08
twenty-four papers selected by
Connor Rogerson
University of Cambridge, MRC Cancer Unit


  1. Development. 2020 Nov 03. pii: dev.188516. [Epub ahead of print]
    Kurup JT, Han Z, Jin W, Kidder BL.
      Heterochromatin, which is a densely packed chromatin state that is transcriptionally silent, is a critical regulator of gene expression. However, it is unclear how the repressive histone modification, H4K20me3, or the histone methyltransferase, SUV420H2, regulate embryonic stem (ES) cell fate by patterning the epigenetic landscape. Here, we report that depletion of SUV420H2 leads to a near complete loss of H4K20me3 genome-wide, dysregulated gene expression, and delayed ES cell differentiation. SUV420H2-bound regions are enriched with repetitive DNA elements, which are de-repressed in SUV420H2 knockout ES cells. Moreover, SUV420H2 regulation of H4K20me3-marked heterochromatin controls chromatin architecture, including fine-scale chromatin interactions in pluripotent ES cells. SUV420H2 plays a critical role in stabilizing the three-dimensional (3D) chromatin landscape of ES cells, where loss of SUV420H2 results in A/B compartment switching, perturbed chromatin insulation, and altered chromatin interactions of pericentric heterochromatin and surrounding regions, indicative of localized decondensation. In addition, depletion of SUV420H2 resulted in compromised interactions between H4K20me3 and gene regulatory regions. Together, these findings describe a novel role for SUV420H2 in regulating the chromatin landscape of ES cells.
    Keywords:  3D genome; Chip-Seq; Chromatin; Chromatin architecture; Embryonic stem cells; Epigenetics; H4K20me3; Pluripotent; SUV420H2
    DOI:  https://doi.org/10.1242/dev.188516
  2. Stem Cells. 2020 Nov 07.
    YuXiu L, Yao X, ChuJiao L, Qian Z, Shu Z, Fei P, Huan L, Zhi C.
      Mouse dental papilla cells (mDPCs) derive from cranial neural crest cells and maintain mesenchymal stem cell characteristics. The differentiation of neural crest cells into odontoblasts is orchestrated by transcription factors regulating the expression of genes whose enhancers are initially inaccessible. However, the identity of the transcription factors driving the emergence of odontoblast lineages remains elusive. In this study, we identified SALL1, a transcription factor that was particularly expressed in pre-odontoblasts, polarizing odontoblasts, and secretory odontoblasts in vivo. Knockdown of Sall1 in mDPCs inhibited their odontoblastic differentiation. In order to identify the regulatory network of Sall1, RNA sequencing and an assay for transposase-accessible chromatin with high-throughput sequencing were performed to analyze the genome-wide direct regulatory targets of SALL1. We found that inhibition of Sall1 expression could decrease the accessibility of some chromatin regions associated with odontoblast lineages at embryonic day 16.5, while these regions remained unaffected at postnatal day 0.5, suggesting that SALL1 regulates the fate of mDPCs by remodeling open chromatin regions at the early bell stage. Specifically, we found that SALL1 could directly increase the accessibility of cis-regulatory elements near Tgf-β2 and within the Runx2 locus. Moreover, co-immunoprecipitation and proximal ligation assays showed that SALL1 could establish functional interactions with RUNX2. Taken together, our results demonstrated that SALL1 positively regulates the commitment of odontoblast lineages by interacting with RUNX2 and directly activating Tgf-β2 at an early stage. © AlphaMed Press 2020 SIGNIFICANCE STATEMENT: Mouse dental papilla cells (mDPCs), considered to be progenitors of odontoblasts, are critical for dentin formation. However, the mechanism by which transcription factors spatially and temporally drive lineage specification in mDPCs is only partially understood. The present study demonstrates that SALL1 positively regulates the commitment of mDPCs to form odontoblastic-like cells at the early differentiation stage. An assay for transposase-accessible chromatin with high-throughput sequencing and RNA sequencing were used to identify Runx2 and Tgf-β2 as direct regulatory targets of SALL1 at embryonic day 16.5. These findings shed light on our understanding of the molecular regulatory mechanism of odontoblastic differentiation, with a possible positive impact on tooth regeneration in the future.
    Keywords:  Differentiation; Epigenetics; Mesenchymal stem cells (MSCs); Transcription factors; Transcriptional regulation
    DOI:  https://doi.org/10.1002/stem.3298
  3. Cell Rep. 2020 Nov 03. pii: S2211-1247(20)31322-X. [Epub ahead of print]33(5): 108333
    Haniuda K, Fukao S, Kitamura D.
      The germinal center (GC) reaction is essential for long-lived humoral immunity. However, molecular requirements for the induction of Bcl6, the master regulator for GC B cell differentiation, remain unclear. Through screening for cytokines and other stimuli that regulate Bcl6 expression, we identify IL-4 as the strongest inducer. IL-4 signaling alters the metabolomic profile in activated B cells and induces accumulation of the TCA cycle intermediate α-ketoglutarate (αKG), which is required for activation of the Bcl6 gene locus. Mechanistically, after IL-4 treatment, STAT6 bound to the known enhancers in the Bcl6 locus recruits UTX, a demethylase for the repressive histone mark H3K27me3 that requires αKG as a cofactor. In turn, the H3K27me3 demethylation activates the enhancers and transcription of the Bcl6 gene. We propose that IL-4-mediated metabolic reprogramming in B cells is pivotal for epigenomic activation of Bcl6 expression to promote GC B cell differentiation.
    Keywords:  B cell; Bcl6; H3K27me3; TCA cycle; differentiation; epigenomic remodeling; germinal center; histone demethylase; metabolism; α-ketoglutarate
    DOI:  https://doi.org/10.1016/j.celrep.2020.108333
  4. Nat Commun. 2020 11 03. 11(1): 5560
    Zhao D, Zhang L, Zhang M, Xia B, Lv J, Gao X, Wang G, Meng Q, Yi Y, Zhu S, Tomoiaga AS, Lee MG, Cooke JP, Cao Q, Chen K.
      Cancers result from a set of genetic and epigenetic alterations. Most known oncogenes were identified by gain-of-function mutations in cancer, yet little is known about their epigenetic features. Through integrative analysis of 11,596 epigenomic profiles and mutations from >8200 tumor-normal pairs, we discover broad genic repression domains (BGRD) on chromatin as an epigenetic signature for oncogenes. A BGRD is a widespread enrichment domain of the repressive histone modification H3K27me3 and is further enriched with multiple other repressive marks including H3K9me3, H3K9me2, and H3K27me2. Further, BGRD displays widespread enrichment of repressed cis-regulatory elements. Shortening of BGRDs is linked to derepression of transcription. BGRDs at oncogenes tend to be conserved across normal cell types. Putative tumor-promoting genes and lncRNAs defined using BGRDs are experimentally verified as required for cancer phenotypes. Therefore, BGRDs play key roles in epigenetic regulation of cancer and provide a direction for mutation-independent discovery of oncogenes.
    DOI:  https://doi.org/10.1038/s41467-020-18913-8
  5. Nat Commun. 2020 Nov 06. 11(1): 5647
    Ni K, Ren J, Xu X, He Y, Finney R, Braun SMG, Hathaway NA, Crabtree GR, Muegge K.
      The human Immunodeficiency Centromeric Instability Facial Anomalies (ICF) 4 syndrome is a severe disease with increased mortality caused by mutation in the LSH gene. Although LSH belongs to a family of chromatin remodeling proteins, it remains unknown how LSH mediates its function on chromatin in vivo. Here, we use chemical-induced proximity to rapidly recruit LSH to an engineered locus and find that LSH specifically induces macroH2A1.2 and macroH2A2 deposition in an ATP-dependent manner. Tethering of LSH induces transcriptional repression and silencing is dependent on macroH2A deposition. Loss of LSH decreases macroH2A enrichment at repeat sequences and results in transcriptional reactivation. Likewise, reduction of macroH2A by siRNA interference mimicks transcriptional reactivation. ChIP-seq analysis confirmed that LSH is a major regulator of genome-wide macroH2A distribution. Tethering of ICF4 mutations fails to induce macroH2A deposition and ICF4 patient cells display reduced macroH2A deposition and transcriptional reactivation supporting a pathogenic role for altered marcoH2A deposition. We propose that LSH is a major chromatin modulator of the histone variant macroH2A and that its ability to insert marcoH2A into chromatin and transcriptionally silence is disturbed in the ICF4 syndrome.
    DOI:  https://doi.org/10.1038/s41467-020-19159-0
  6. Development. 2020 Nov 03. pii: dev.190298. [Epub ahead of print]
    Yi S, Huang X, Zhou S, Zhou Y, Anderson MK, Zúñiga-Pflücker JC, Luan Q, Li Y.
      E proteins transcription factors are critical for many cell fate decisions. However, the roles of E proteins in the germ-layer specification of human embryonic stem cells (hESC) are poorly understood. We disrupted the TCF3 gene locus to delete the E protein E2A in hESCs. E2A KO hESCs retained key features of pluripotency, but displayed decreased neural ectoderm coupled with enhanced mesoendoderm outcomes. Genome-wide analyses showed that E2A directly regulates neural ectoderm and Nodal pathway genes. Accordingly, inhibition of Nodal or E2A overexpression partially rescued the neural ectoderm defect in E2A KO hESCs. Loss of E2A had little impact on the epigenetic landscape of hESCs, whereas E2A KO neural precursors displayed increased accessibility of the gene locus encoding the Nodal agonist CRIPTO. Double-deletion of both E2A and HEB (TCF12) resulted in a more severe neural ectoderm defect. Therefore, this study reveals critical context-dependent functions for E2A in human neural ectoderm fate-specification.
    Keywords:  E2A; Human embryonic stem cells; Neural differentiation; Nodal signaling pathway; PRC2 complex
    DOI:  https://doi.org/10.1242/dev.190298
  7. Cancer Res. 2020 Nov 06. pii: canres.3922.2019. [Epub ahead of print]
    Tomihara H, Carbone F, Perelli L, Huang JK, Soeung M, Rose JL, Robinson FS, Lissanu Deribe Y, Feng N, Takeda M, Inoue A, Deem AK, Maitra A, Msaouel P, Tannir NM, Draetta GF, Bristow CA, Carugo A, Genovese G.
      Cellular de-differentiation is a key mechanism driving cancer progression. Acquisition of mesenchymal features has been associated with drug resistance, poor prognosis, and disease relapse in many tumor types. Therefore, successful targeting of tumors harboring these characteristics is a priority in oncology practice. The SWItch/Sucrose Non-Fermentable (SWI/SNF) chromatin remodeling complex has also emerged as a critical player in tumor progression, leading to the identification of several SWI/SNF complex genes as potential disease biomarkers and targets of anti-cancer therapies. AT-rich interaction domain-containing protein 1A (ARID1A) is a component of SWI/SNF, and mutations in ARID1A represent one of the most frequent molecular alterations in human cancers. ARID1A mutations occur in ~10% of pancreatic ductal adenocarcinomas (PDAC), but whether these mutations confer a therapeutic opportunity remains unclear. Here we demonstrate that loss of ARID1A promotes an epithelial-mesenchymal transition (EMT) phenotype and sensitizes PDAC cells to a clinical inhibitor of HSP90, NVP-AUY922, both in vitro and in vivo. While loss of ARID1A alone did not significantly affect proliferative potential or rate of apoptosis, ARID1A-deficient cells were sensitized to HSP90 inhibition, potentially by promoting the degradation of intermediate filaments driving EMT, resulting in cell death. Our results describe a mechanistic link between ARID1A defects and a quasi-mesenchymal phenotype, suggesting that deleterious mutations in ARID1A associated with protein loss exhibits potential as a biomarker for PDAC patients who may benefit by HSP90-targeting drugs treatment.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-19-3922
  8. PLoS Comput Biol. 2020 Nov 02. 16(11): e1008334
    Chen L, Capra JA.
      Deep neural networks (DNNs) have achieved state-of-the-art performance in identifying gene regulatory sequences, but they have provided limited insight into the biology of regulatory elements due to the difficulty of interpreting the complex features they learn. Several models of how combinatorial binding of transcription factors, i.e. the regulatory grammar, drives enhancer activity have been proposed, ranging from the flexible TF billboard model to the stringent enhanceosome model. However, there is limited knowledge of the prevalence of these (or other) sequence architectures across enhancers. Here we perform several hypothesis-driven analyses to explore the ability of DNNs to learn the regulatory grammar of enhancers. We created synthetic datasets based on existing hypotheses about combinatorial transcription factor binding site (TFBS) patterns, including homotypic clusters, heterotypic clusters, and enhanceosomes, from real TF binding motifs from diverse TF families. We then trained deep residual neural networks (ResNets) to model the sequences under a range of scenarios that reflect real-world multi-label regulatory sequence prediction tasks. We developed a gradient-based unsupervised clustering method to extract the patterns learned by the ResNet models. We demonstrated that simulated regulatory grammars are best learned in the penultimate layer of the ResNets, and the proposed method can accurately retrieve the regulatory grammar even when there is heterogeneity in the enhancer categories and a large fraction of TFBS outside of the regulatory grammar. However, we also identify common scenarios where ResNets fail to learn simulated regulatory grammars. Finally, we applied the proposed method to mouse developmental enhancers and were able to identify the components of a known heterotypic TF cluster. Our results provide a framework for interpreting the regulatory rules learned by ResNets, and they demonstrate that the ability and efficiency of ResNets in learning the regulatory grammar depends on the nature of the prediction task.
    DOI:  https://doi.org/10.1371/journal.pcbi.1008334
  9. J Biol Chem. 2020 Nov 03. pii: jbc.RA120.014598. [Epub ahead of print]
    Wu L, Zhao G, Xu S, Kuang J, Min J, Wu G, Wang T, Wang B, Zhu P, Pei D, Liu J.
      Somatic cells can be reprogrammed into pluripotent stem cells with a minimal set of defined factors, Oct3/4, Sox2 and Klf4, also known as OSK,though this reprogramming is somewhat inefficient. Recent work has identified other nuclear factors, including SALL4, that can synergize with the OSK factors to improve reprogramming dynamics, [CG1] but the specific role of each of these factors remains poorly understood..by a set of defined factors. However, the reprogramming ability and underlying mechanism for each factor remains poorly understood. Here, we show Cecr2 as a target of SALL4 in accelerating OSK induced reprogramming. By screening a group of putative downstream targets, we identified CECR2, a histone acetyl-lysine reader, can significantly promote OKS induced reprogramming as a SALL4 effector. Mechanically, SALL4 activates Cecr2 by directly binging to its promotor region and CECR2 in turn promotes reprogramming through forming a SMARCA1-contained chromatin remodeling complex with its DTT domain. Our findings suggest that CECR2 is a novel reprogramming factors and it works through a protein network to overcome epigenetic barriers during reprogramming.
    Keywords:  cell biology; chromatin structure; epigenetics; reprogramming; transcription factor
    DOI:  https://doi.org/10.1074/jbc.RA120.014598
  10. Sci Adv. 2020 Nov;pii: eaaz1410. [Epub ahead of print]6(45):
    Sun J, Feng H, Xing W, Han Y, Suo J, Yallowitz AR, Qian N, Shi Y, Greenblatt MB, Zou W.
      Bone fracture is repaired predominantly through endochondral ossification. However, the regulation of endochondral ossification by key factors during fracture healing remains largely enigmatic. Here, we identify histone modification enzyme LSD1 as a critical factor regulating endochondral ossification during bone regeneration. Loss of LSD1 in Prx1 lineage cells severely impaired bone fracture healing. Mechanistically, LSD1 tightly controls retinoic acid signaling through regulation of Aldh1a2 expression level. The increased retinoic acid signaling in LSD1-deficient mice suppressed SOX9 expression and impeded the cartilaginous callus formation during fracture repair. The discovery that LSD1 can regulate endochondral ossification during fracture healing will benefit the understanding of bone regeneration and have implications for regenerative medicine.
    DOI:  https://doi.org/10.1126/sciadv.aaz1410
  11. Nat Genet. 2020 Nov 02.
    Fan H, Lu J, Guo Y, Li D, Zhang ZM, Tsai YH, Pi WC, Ahn JH, Gong W, Xiang Y, Allison DF, Geng H, He S, Diao Y, Chen WY, Strahl BD, Cai L, Song J, Wang GG.
      Trimethylated histone H3 lysine 27 (H3K27me3) regulates gene repression, cell-fate determination and differentiation. We report that a conserved bromo-adjacent homology (BAH) module of BAHCC1 (BAHCC1BAH) 'recognizes' H3K27me3 specifically and enforces silencing of H3K27me3-demarcated genes in mammalian cells. Biochemical, structural and integrated chromatin immunoprecipitation-sequencing-based analyses demonstrate that direct readout of H3K27me3 by BAHCC1 is achieved through a hydrophobic trimethyl-L-lysine-binding 'cage' formed by BAHCC1BAH, mediating colocalization of BAHCC1 and H3K27me3-marked genes. BAHCC1 is highly expressed in human acute leukemia and interacts with transcriptional corepressors. In leukemia, depletion of BAHCC1, or disruption of the BAHCC1BAH-H3K27me3 interaction, causes derepression of H3K27me3-targeted genes that are involved in tumor suppression and cell differentiation, leading to suppression of oncogenesis. In mice, introduction of a germline mutation at Bahcc1 to disrupt its H3K27me3 engagement causes partial postnatal lethality, supporting a role in development. This study identifies an H3K27me3-directed transduction pathway in mammals that relies on a conserved BAH 'reader'.
    DOI:  https://doi.org/10.1038/s41588-020-00729-3
  12. Nucleic Acids Res. 2020 Nov 05. pii: gkaa995. [Epub ahead of print]
    Fang L, Li Y, Ma L, Xu Q, Tan F, Chen G.
      Gene regulatory networks (GRNs) formed by transcription factors (TFs) and their downstream target genes play essential roles in gene expression regulation. Moreover, GRNs can be dynamic changing across different conditions, which are crucial for understanding the underlying mechanisms of disease pathogenesis. However, no existing database provides comprehensive GRN information for various human and mouse normal tissues and diseases at the single-cell level. Based on the known TF-target relationships and the large-scale single-cell RNA-seq data collected from public databases as well as the bulk data of The Cancer Genome Atlas and the Genotype-Tissue Expression project, we systematically predicted the GRNs of 184 different physiological and pathological conditions of human and mouse involving >633 000 cells and >27 700 bulk samples. We further developed GRNdb, a freely accessible and user-friendly database (http://www.grndb.com/) for searching, comparing, browsing, visualizing, and downloading the predicted information of 77 746 GRNs, 19 687 841 TF-target pairs, and related binding motifs at single-cell/bulk resolution. GRNdb also allows users to explore the gene expression profile, correlations, and the associations between expression levels and the patient survival of diverse cancers. Overall, GRNdb provides a valuable and timely resource to the scientific community to elucidate the functions and mechanisms of gene expression regulation in various conditions.
    DOI:  https://doi.org/10.1093/nar/gkaa995
  13. Nat Commun. 2020 11 05. 11(1): 5609
    Seif E, Kang JJ, Sasseville C, Senkovich O, Kaltashov A, Boulier EL, Kapur I, Kim CA, Francis NJ.
      Polycomb Group (PcG) proteins organize chromatin at multiple scales to regulate gene expression. A conserved Sterile Alpha Motif (SAM) in the Polycomb Repressive Complex 1 (PRC1) subunit Polyhomeotic (Ph) has been shown to play an important role in chromatin compaction and large-scale chromatin organization. Ph SAM forms helical head to tail polymers, and SAM-SAM interactions between chromatin-bound Ph/PRC1 are believed to compact chromatin and mediate long-range interactions. To understand the underlying mechanism, here we analyze the effects of Ph SAM on chromatin in vitro. We find that incubation of chromatin or DNA with a truncated Ph protein containing the SAM results in formation of concentrated, phase-separated condensates. Ph SAM-dependent condensates can recruit PRC1 from extracts and enhance PRC1 ubiquitin ligase activity towards histone H2A. We show that overexpression of Ph with an intact SAM increases ubiquitylated H2A in cells. Thus, SAM-induced phase separation, in the context of Ph, can mediate large-scale compaction of chromatin into biochemical compartments that facilitate histone modification.
    DOI:  https://doi.org/10.1038/s41467-020-19435-z
  14. J Immunol. 2020 Nov 02. pii: ji2000537. [Epub ahead of print]
    Cameron J, Martino P, Nguyen L, Li X.
      Transcription factor Foxp3 specifies and maintains regulatory T cell (Treg) identity. During Treg differentiation, a CpG-rich Foxp3 intronic enhancer, conserved noncoding sequence 2 (CNS2), is activated via DNA demethylation to establish epigenetic memory of Foxp3 expression to protect Treg identity. However, it is unclear how this epigenetic memory of Foxp3 expression is established, as CNS2 is thought to be demethylated independently of Foxp3 expression. In this article, we uncover an unexpected causal relationship between Foxp3-transcriptional activation and CNS2 demethylation in mice. CRISPR/dCas9-mediated Foxp3-transcriptional activation elicits CNS2 demethylation. Sustaining Foxp3-transcriptional activation in induced Tregs also promotes CNS2 demethylation, enhancing Treg lineage stability and suppressive function. Importantly, CRISPR-mediated silencing of Foxp3 transcription, but not protein expression, abolishes CNS2 demethylation. The novel finding that Foxp3-transcriptional activation promotes CNS2 demethylation may facilitate the development of Treg-based therapies and represent a general mechanism for the establishment of epigenetic memory of immune gene expression.
    DOI:  https://doi.org/10.4049/jimmunol.2000537
  15. Nat Commun. 2020 11 05. 11(1): 5612
    Nora EP, Caccianini L, Fudenberg G, So K, Kameswaran V, Nagle A, Uebersohn A, Hajj B, Saux AL, Coulon A, Mirny LA, Pollard KS, Dahan M, Bruneau BG.
      Current models propose that boundaries of mammalian topologically associating domains (TADs) arise from the ability of the CTCF protein to stop extrusion of chromatin loops by cohesin. While the orientation of CTCF motifs determines which pairs of CTCF sites preferentially stabilize loops, the molecular basis of this polarity remains unclear. By combining ChIP-seq and single molecule live imaging we report that CTCF positions cohesin, but does not control its overall binding dynamics on chromatin. Using an inducible complementation system, we find that CTCF mutants lacking the N-terminus cannot insulate TADs properly. Cohesin remains at CTCF sites in this mutant, albeit with reduced enrichment. Given the orientation of CTCF motifs presents the N-terminus towards cohesin as it translocates from the interior of TADs, these observations explain how the orientation of CTCF binding sites translates into genome folding patterns.
    DOI:  https://doi.org/10.1038/s41467-020-19283-x
  16. BMC Genomics. 2020 Nov 02. 21(1): 754
    Sethi S, Vorontsov IE, Kulakovskiy IV, Greenaway S, Williams J, Makeev VJ, Brown SDM, Simon MM, Mallon AM.
      BACKGROUND: Efforts to elucidate the function of enhancers in vivo are underway but their vast numbers alongside differing enhancer architectures make it difficult to determine their impact on gene activity. By systematically annotating multiple mouse tissues with super- and typical-enhancers, we have explored their relationship with gene function and phenotype.RESULTS: Though super-enhancers drive high total- and tissue-specific expression of their associated genes, we find that typical-enhancers also contribute heavily to the tissue-specific expression landscape on account of their large numbers in the genome. Unexpectedly, we demonstrate that both enhancer types are preferentially associated with relevant 'tissue-type' phenotypes and exhibit no difference in phenotype effect size or pleiotropy. Modelling regulatory data alongside molecular data, we built a predictive model to infer gene-phenotype associations and use this model to predict potentially novel disease-associated genes.
    CONCLUSION: Overall our findings reveal that differing enhancer architectures have a similar impact on mammalian phenotypes whilst harbouring differing cellular and expression effects. Together, our results systematically characterise enhancers with predicted phenotypic traits endorsing the role for both types of enhancers in human disease and disorders.
    Keywords:  Expression; Gene-phenotype prediction; Phenotypes; Protein-protein interactions; Super-enhancers; Tissue-specificity; Transcription factors; Typical-enhancers
    DOI:  https://doi.org/10.1186/s12864-020-07109-5
  17. Nat Commun. 2020 11 04. 11(1): 5581
    Hauberg ME, Creus-Muncunill J, Bendl J, Kozlenkov A, Zeng B, Corwin C, Chowdhury S, Kranz H, Hurd YL, Wegner M, Børglum AD, Dracheva S, Ehrlich ME, Fullard JF, Roussos P.
      The chromatin landscape of human brain cells encompasses key information to understanding brain function. Here we use ATAC-seq to profile the chromatin structure in four distinct populations of cells (glutamatergic neurons, GABAergic neurons, oligodendrocytes, and microglia/astrocytes) from three different brain regions (anterior cingulate cortex, dorsolateral prefrontal cortex, and primary visual cortex) in human postmortem brain samples. We find that chromatin accessibility varies greatly by cell type and, more moderately, by brain region, with glutamatergic neurons showing the largest regional variability. Transcription factor footprinting implicates cell-specific transcriptional regulators and infers cell-specific regulation of protein-coding genes, long intergenic noncoding RNAs and microRNAs. In vivo transgenic mouse experiments validate the cell type specificity of several of these human-derived regulatory sequences. We find that open chromatin regions in glutamatergic neurons are enriched for neuropsychiatric risk variants, particularly those associated with schizophrenia. Integration of cell-specific chromatin data with a bulk tissue study of schizophrenia brains increases statistical power and confirms that glutamatergic neurons are most affected. These findings illustrate the utility of studying the cell-type-specific epigenome in complex tissues like the human brain, and the potential of such approaches to better understand the genetic basis of human brain function.
    DOI:  https://doi.org/10.1038/s41467-020-19319-2
  18. Sci Rep. 2020 Nov 03. 10(1): 18902
    Vardaka P, Lozano T, Bot C, Ellery J, Whiteside SK, Imianowski CJ, Farrow S, Walker S, Okkenhaug H, Yang J, Okkenhaug K, Kuo P, Roychoudhuri R.
      Whereas effector CD4+ and CD8+ T cells promote immune activation and can drive clearance of infections and cancer, CD4+ regulatory T (Treg) cells suppress their function, contributing to both immune homeostasis and cancer immunosuppression. The transcription factor BACH2 functions as a pervasive regulator of T cell differentiation, promoting development of CD4+ Treg cells and suppressing the effector functions of multiple effector T cell (Teff) lineages. Here, we report the development of a stable cell-based bioluminescence assay of the transcription factor activity of BACH2. Tetracycline-inducible BACH2 expression resulted in suppression of phorbol 12-myristate 13-acetate (PMA)/ionomycin-driven activation of a luciferase reporter containing BACH2/AP-1 target sequences from the mouse Ifng + 18k enhancer. BACH2 expression repressed the luciferase signal in a dose-dependent manner but this activity was abolished at high levels of AP-1 signalling, suggesting contextual regulation of AP-1 driven gene expression by BACH2. Finally, using the reporter assay developed, we find that the histone deacetylase 3 (HDAC3)-selective inhibitor, RGFP966, inhibits BACH2-mediated repression of signal-driven luciferase expression. In addition to enabling mechanistic studies, this cell-based reporter may enable identification of small molecule agonists or antagonists of BACH2 function for drug development.
    DOI:  https://doi.org/10.1038/s41598-020-75732-z
  19. Genome Biol. 2020 Nov 03. 21(1): 269
    Fang K, Huang W, Sun YM, Chen TQ, Zeng ZC, Yang QQ, Pan Q, Han C, Sun LY, Luo XQ, Wang WT, Chen YQ.
      BACKGROUND: Long noncoding enhancer RNAs (lnc-eRNAs) are a subset of stable eRNAs identified from annotated lncRNAs. They might act as enhancer activity-related therapeutic targets in cancer. However, the underlying mechanism of epigenetic activation and their function in cancer initiation and progression remain largely unknown.RESULTS: We identify a set of lncRNAs as lnc-eRNAs according to the epigenetic signatures of enhancers. We show that these lnc-eRNAs are broadly activated in MLL-rearranged leukemia (MLL leukemia), an aggressive leukemia caused by a chromosomal translocation, through a mechanism by which the HOXA cluster initiates enhancer activity, and the epigenetic reader BRD4 cooperates with the coregulator MLL fusion oncoprotein to induce transcriptional activation. To demonstrate the functional roles of lnc-eRNAs, two newly identified lnc-eRNAs transcribed from the SEELA eRNA cluster (SEELA), SEELA1 and SEELA2, are chosen for further studies. The results show that SEELA mediated cis-activated transcription of the nearby oncogene Serine incorporate 2 (SERINC2) by directly binding to the K31 amino acid (aa) of histone H4. Chromatin-bound SEELA strengthens the interaction between chromatin and histone modifiers to promote histone recognition and oncogene transcription. Further studies show that the SEELA-SERINC2 axis regulated aspects of cancer metabolism, such as sphingolipid synthesis, to affect leukemia progression.
    CONCLUSIONS: This study shows that lnc-eRNAs are epigenetically activated by cancer-initiating oncoproteins and uncovers a cis-activating mechanism of oncogene transcription control based on lnc-eRNA-mediated epigenetic regulation of enhancer activity, providing insights into the critical roles of lnc-eRNAs in cancer initiation and progression.
    Keywords:  Enhancer activity; Histone H4; Histone recognition; Lnc-eRNA; MLL leukemia; SEELA; Sphingolipid metabolism
    DOI:  https://doi.org/10.1186/s13059-020-02186-x
  20. Cell Metab. 2020 Nov 03. pii: S1550-4131(20)30538-6. [Epub ahead of print]32(5): 889-900.e7
    Ludikhuize MC, Meerlo M, Gallego MP, Xanthakis D, Burgaya Julià M, Nguyen NTB, Brombacher EC, Liv N, Maurice MM, Paik JH, Burgering BMT, Rodriguez Colman MJ.
      Differential WNT and Notch signaling regulates differentiation of Lgr5+ crypt-based columnar cells (CBCs) into intestinal cell lineages. Recently we showed that mitochondrial activity supports CBCs, while adjacent Paneth cells (PCs) show reduced mitochondrial activity. This implies that CBC differentiation into PCs involves a metabolic transition toward downregulation of mitochondrial dependency. Here we show that Forkhead box O (FoxO) transcription factors and Notch signaling interact in determining CBC fate. In agreement with the organoid data, Foxo1/3/4 deletion in mouse intestine induces secretory cell differentiation. Importantly, we show that FOXO and Notch signaling converge on regulation of mitochondrial fission, which in turn provokes stem cell differentiation into goblet cells and PCs. Finally, scRNA-seq-based reconstruction of CBC differentiation trajectories supports the role of FOXO, Notch, and mitochondria in secretory differentiation. Together, this points at a new signaling-metabolic axis in CBC differentiation and highlights the importance of mitochondria in determining stem cell fate.
    Keywords:  FOXO; Notch; differentiation; intestine; metabolism; mitochondria; stem cells
    DOI:  https://doi.org/10.1016/j.cmet.2020.10.005
  21. Nat Commun. 2020 11 03. 11(1): 5549
    Cyrta J, Augspach A, De Filippo MR, Prandi D, Thienger P, Benelli M, Cooley V, Bareja R, Wilkes D, Chae SS, Cavaliere P, Dephoure N, Uldry AC, Lagache SB, Roma L, Cohen S, Jaquet M, Brandt LP, Alshalalfa M, Puca L, Sboner A, Feng F, Wang S, Beltran H, Lotan T, Spahn M, Kruithof-de Julio M, Chen Y, Ballman KV, Demichelis F, Piscuoglio S, Rubin MA.
      Advanced prostate cancer initially responds to hormonal treatment, but ultimately becomes resistant and requires more potent therapies. One mechanism of resistance observed in around 10-20% of these patients is lineage plasticity, which manifests in a partial or complete small cell or neuroendocrine prostate cancer (NEPC) phenotype. Here, we investigate the role of the mammalian SWI/SNF (mSWI/SNF) chromatin remodeling complex in NEPC. Using large patient datasets, patient-derived organoids and cancer cell lines, we identify mSWI/SNF subunits that are deregulated in NEPC and demonstrate that SMARCA4 (BRG1) overexpression is associated with aggressive disease. We also show that SWI/SNF complexes interact with different lineage-specific factors in NEPC compared to prostate adenocarcinoma. These data point to a role for mSWI/SNF complexes in therapy-related lineage plasticity, which may also be relevant for other solid tumors.
    DOI:  https://doi.org/10.1038/s41467-020-19328-1
  22. PLoS One. 2020 ;15(11): e0241698
    Yu B, Doni Jayavelu N, Battle SL, Mar JC, Schimmel T, Cohen J, Hawkins RD.
      Oocyte maturation is a coordinated process that is tightly linked to reproductive potential. A better understanding of gene regulation during human oocyte maturation will not only answer an important question in biology, but also facilitate the development of in vitro maturation technology as a fertility treatment. We generated single-cell transcriptome and used our previously published single-cell methylome data from human oocytes at different maturation stages to investigate how genes are regulated during oocyte maturation, focusing on the potential regulatory role of non-CpG methylation. DNMT3B, a gene encoding a key non-CpG methylation enzyme, is one of the 1,077 genes upregulated in mature oocytes, which may be at least partially responsible for the increased non-CpG methylation as oocytes mature. Non-CpG differentially methylated regions (DMRs) between mature and immature oocytes have multiple binding motifs for transcription factors, some of which bind with DNMT3B and may be important regulators of oocyte maturation through non-CpG methylation. Over 98% of non-CpG DMRs locate in transposable elements, and these DMRs are correlated with expression changes of the nearby genes. Taken together, this data indicates that global non-CpG hypermethylation during oocyte maturation may play an active role in gene expression regulation, potentially through the interaction with transcription factors.
    DOI:  https://doi.org/10.1371/journal.pone.0241698
  23. Nat Commun. 2020 11 05. 11(1): 5596
    Wang Y, Deng P, Liu Y, Wu Y, Chen Y, Guo Y, Zhang S, Zheng X, Zhou L, Liu W, Li Q, Lin W, Qi X, Ou G, Wang C, Yuan Q.
      Age-related osteoporosis is characterized by the deterioration in bone volume and strength, partly due to the dysfunction of bone marrow mesenchymal stromal/stem cells (MSCs) during aging. Alpha-ketoglutarate (αKG) is an essential intermediate in the tricarboxylic acid (TCA) cycle. Studies have revealed that αKG extends the lifespan of worms and maintains the pluripotency of embryonic stem cells (ESCs). Here, we show that the administration of αKG increases the bone mass of aged mice, attenuates age-related bone loss, and accelerates bone regeneration of aged rodents. αKG ameliorates the senescence-associated (SA) phenotypes of bone marrow MSCs derived from aged mice, as well as promoting their proliferation, colony formation, migration, and osteogenic potential. Mechanistically, αKG decreases the accumulations of H3K9me3 and H3K27me3, and subsequently upregulates BMP signaling and Nanog expression. Collectively, our findings illuminate the role of αKG in rejuvenating MSCs and ameliorating age-related osteoporosis, with a promising therapeutic potential in age-related diseases.
    DOI:  https://doi.org/10.1038/s41467-020-19360-1
  24. Bioinformatics. 2020 Nov 02. pii: btaa927. [Epub ahead of print]
    Liu L, Chandrashekar P, Zeng B, Sanderford MD, Kumar S, Gibson G.
      MOTIVATION: Expression quantitative trait loci (eQTL) harbor genetic variants modulating gene transcription. Fine mapping of regulatory variants at these loci is a daunting task due to the juxtaposition of causal and linked variants at a locus as well as the likelihood of interactions among multiple variants. This problem is exacerbated in genes with multiple cis-acting eQTL, where superimposed effects of adjacent loci further distort the association signals.RESULTS: We developed a novel algorithm, TreeMap, that identifies putative causal variants in cis-eQTL accounting for multisite effects and genetic linkage at a locus. Guided by the hierarchical structure of linkage disequilibrium, TreeMap performs an organized search for individual and multiple causal variants. Via extensive simulations, we show that TreeMap detects co-regulating variants more accurately than current methods. Furthermore, its high computational efficiency enables genome-wide analysis of long-range eQTL. We applied TreeMap to GTEx data of brain hippocampus samples and transverse colon samples to search for eQTL in gene bodies and in 4 Mbps gene-flanking regions, discovering numerous distal eQTL. Furthermore, we found concordant distal eQTL that were present in both brain and colon samples, implying long-range regulation of gene expression.
    AVAILABILITY: TreeMap is available as an R package enabled for parallel processing at https://github.com/liliulab/treemap.
    CONTACT: liliu@asu.edu, s.kumar@temple.edu, greg.gibson@biology.gatech.edu.
    SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
    DOI:  https://doi.org/10.1093/bioinformatics/btaa927