bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2021‒04‒18
thirty-one papers selected by
Connor Rogerson
University of Cambridge, MRC Cancer Unit


  1. Genome Biol. 2021 Apr 15. 22(1): 105
      Muscle-invasive bladder cancers are characterized by their distinct expression of luminal and basal genes, which could be used to predict key clinical features such as disease progression and overall survival. Transcriptionally, FOXA1, GATA3, and PPARG are shown to be essential for luminal subtype-specific gene regulation and subtype switching, while TP63, STAT3, and TFAP2 family members are critical for regulation of basal subtype-specific genes. Despite these advances, the underlying epigenetic mechanisms and 3D chromatin architecture responsible for subtype-specific regulation in bladder cancer remain unknown. RESULT: We determine the genome-wide transcriptome, enhancer landscape, and transcription factor binding profiles of FOXA1 and GATA3 in luminal and basal subtypes of bladder cancer. Furthermore, we report the first-ever mapping of genome-wide chromatin interactions by Hi-C in both bladder cancer cell lines and primary patient tumors. We show that subtype-specific transcription is accompanied by specific open chromatin and epigenomic marks, at least partially driven by distinct transcription factor binding at distal enhancers of luminal and basal bladder cancers. Finally, we identify a novel clinically relevant transcription factor, Neuronal PAS Domain Protein 2 (NPAS2), in luminal bladder cancers that regulates other subtype-specific genes and influences cancer cell proliferation and migration. CONCLUSION: In summary, our work identifies unique epigenomic signatures and 3D genome structures in luminal and basal urinary bladder cancers and suggests a novel link between the circadian transcription factor NPAS2 and a clinical bladder cancer subtype.
    DOI:  https://doi.org/10.1186/s13059-021-02325-y
  2. Nat Commun. 2021 04 15. 12(1): 2277
      Determining the epigenetic program that generates unique cell types in the kidney is critical for understanding cell-type heterogeneity during tissue homeostasis and injury response. Here, we profile open chromatin and gene expression in developing and adult mouse kidneys at single cell resolution. We show critical reliance of gene expression on distal regulatory elements (enhancers). We reveal key cell type-specific transcription factors and major gene-regulatory circuits for kidney cells. Dynamic chromatin and expression changes during nephron progenitor differentiation demonstrates that podocyte commitment occurs early and is associated with sustained Foxl1 expression. Renal tubule cells follow a more complex differentiation, where Hfn4a is associated with proximal and Tfap2b with distal fate. Mapping single nucleotide variants associated with human kidney disease implicates critical cell types, developmental stages, genes, and regulatory mechanisms. The single cell multi-omics atlas reveals key chromatin remodeling events and gene expression dynamics associated with kidney development.
    DOI:  https://doi.org/10.1038/s41467-021-22266-1
  3. Nat Biotechnol. 2021 Apr 12.
      Methods for quantifying gene expression1 and chromatin accessibility2 in single cells are well established, but single-cell analysis of chromatin regions with specific histone modifications has been technically challenging. In this study, we adapted the CUT&Tag method3 to scalable nanowell and droplet-based single-cell platforms to profile chromatin landscapes in single cells (scCUT&Tag) from complex tissues and during the differentiation of human embryonic stem cells. We focused on profiling polycomb group (PcG) silenced regions marked by histone H3 Lys27 trimethylation (H3K27me3) in single cells as an orthogonal approach to chromatin accessibility for identifying cell states. We show that scCUT&Tag profiling of H3K27me3 distinguishes cell types in human blood and allows the generation of cell-type-specific PcG landscapes from heterogeneous tissues. Furthermore, we used scCUT&Tag to profile H3K27me3 in a patient with a brain tumor before and after treatment, identifying cell types in the tumor microenvironment and heterogeneity in PcG activity in the primary sample and after treatment.
    DOI:  https://doi.org/10.1038/s41587-021-00865-z
  4. Cell Rep. 2021 Apr 13. pii: S2211-1247(21)00292-8. [Epub ahead of print]35(2): 108978
      The human fallopian tube harbors the cell of origin for the majority of high-grade serous "ovarian" cancers (HGSCs), but its cellular composition, particularly the epithelial component, is poorly characterized. We perform single-cell transcriptomic profiling of around 53,000 individual cells from 12 primary fallopian specimens to map their major cell types. We identify 10 epithelial subpopulations with diverse transcriptional programs. Based on transcriptional signatures, we reconstruct a trajectory whereby secretory cells differentiate into ciliated cells via a RUNX3high intermediate. Computational deconvolution of advanced HGSCs identifies the "early secretory" population as a likely precursor state for the majority of HGSCs. Its signature comprises both epithelial and mesenchymal features and is enriched in mesenchymal-type HGSCs (p = 6.7 × 10-27), a group known to have particularly poor prognoses. This cellular and molecular compendium of the human fallopian tube in cancer-free women is expected to advance our understanding of the earliest stages of fallopian epithelial neoplasia.
    Keywords:  PAX8; RUNX3; SOX17; ciliated epithelial cells; fallopian tube; microenvironment; ovarian cancer; scRNA-seq; secretory epithelial cells; transcription factor
    DOI:  https://doi.org/10.1016/j.celrep.2021.108978
  5. Nucleic Acids Res. 2021 Apr 13. pii: gkab252. [Epub ahead of print]
      Castration-resistant prostate cancer (CRPC) is a terminal disease and the molecular underpinnings of CRPC development need to be better understood in order to improve its treatment. Here, we report that a transcription factor Yin Yang 1 (YY1) is significantly overexpressed during prostate cancer progression. Functional and cistrome studies of YY1 uncover its roles in promoting prostate oncogenesis in vitro and in vivo, as well as sustaining tumor metabolism including the Warburg effect and mitochondria respiration. Additionally, our integrated genomics and interactome profiling in prostate tumor show that YY1 and bromodomain-containing proteins (BRD2/4) co-occupy a majority of gene-regulatory elements, coactivating downstream targets. Via gene loss-of-function and rescue studies and mutagenesis of YY1-bound cis-elements, we unveil an oncogenic pathway in which YY1 directly binds and activates PFKP, a gene encoding the rate-limiting enzyme for glycolysis, significantly contributing to the YY1-enforced Warburg effect and malignant growth. Altogether, this study supports a master regulator role for YY1 in prostate tumorigenesis and reveals a YY1:BRD2/4-PFKP axis operating in advanced prostate cancer with implications for therapy.
    DOI:  https://doi.org/10.1093/nar/gkab252
  6. Nat Commun. 2021 04 13. 12(1): 2190
      The integration of single cell transcriptome and chromatin accessibility datasets enables a deeper understanding of cell heterogeneity. We performed single nucleus ATAC (snATAC-seq) and RNA (snRNA-seq) sequencing to generate paired, cell-type-specific chromatin accessibility and transcriptional profiles of the adult human kidney. We demonstrate that snATAC-seq is comparable to snRNA-seq in the assignment of cell identity and can further refine our understanding of functional heterogeneity in the nephron. The majority of differentially accessible chromatin regions are localized to promoters and a significant proportion are closely associated with differentially expressed genes. Cell-type-specific enrichment of transcription factor binding motifs implicates the activation of NF-κB that promotes VCAM1 expression and drives transition between a subpopulation of proximal tubule epithelial cells. Our multi-omics approach improves the ability to detect unique cell states within the kidney and redefines cellular heterogeneity in the proximal tubule and thick ascending limb.
    DOI:  https://doi.org/10.1038/s41467-021-22368-w
  7. Nucleic Acids Res. 2021 Apr 13. pii: gkab209. [Epub ahead of print]
      Although overexpression of EZH2, a catalytic subunit of the polycomb repressive complex 2 (PRC2), is an eminent feature of various cancers, the regulation of its abundance and function remains insufficiently understood. We report here that the PRC2 complex is physically associated with ubiquitin-specific protease USP7 in cancer cells where USP7 acts to deubiquitinate and stabilize EZH2. Interestingly, we found that USP7-catalyzed H2BK120ub1 deubiquitination is a prerequisite for chromatin loading of PRC2 thus H3K27 trimethylation, and this process is not affected by H2AK119 ubiquitination catalyzed by PRC1. Genome-wide analysis of the transcriptional targets of the USP7/PRC2 complex identified a cohort of genes including FOXO1 that are involved in cell growth and proliferation. We demonstrated that the USP7/PRC2 complex drives cancer cell proliferation and tumorigenesis in vitro and in vivo. We showed that the expression of both USP7 and EZH2 elevates during tumor progression, corresponding to a diminished FOXO1 expression, and the level of the expression of USP7 and EZH2 strongly correlates with histological grades and prognosis of tumor patients. These results reveal a dual role for USP7 in the regulation of the abundance and function of EZH2, supporting the pursuit of USP7 as a therapeutic target for cancer intervention.
    DOI:  https://doi.org/10.1093/nar/gkab209
  8. Nat Commun. 2021 04 13. 12(1): 2208
      The B1 and B2 lineages of B cells contribute to protection from pathogens in distinct ways. The role of the DNA CpG methylome in specifying these two B-cell fates is still unclear. Here we profile the CpG modifications and transcriptomes of peritoneal B1a and follicular B2 cells, as well as their respective proB cell precursors in the fetal liver and adult bone marrow from wild-type and CD19-Cre Dnmt3a floxed mice lacking DNMT3A in the B lineage. We show that an underlying foundational CpG methylome is stably established during B lineage commitment and is overlaid with a DNMT3A-maintained dynamic methylome that is sculpted in distinct ways in B1a and B2 cells. This dynamic DNMT3A-maintained methylome is composed of novel enhancers that are closely linked to lineage-specific genes. While DNMT3A maintains the methylation state of these enhancers in both B1a and B2 cells, the dynamic methylome undergoes a prominent programmed demethylation event during B1a but not B2 cell development. We propose that the methylation pattern of DNMT3A-maintained enhancers is determined by the coincident recruitment of DNMT3A and TET enzymes, which regulate the developmental expression of B1a and B2 lineage-specific genes.
    DOI:  https://doi.org/10.1038/s41467-021-22458-9
  9. Nat Biotechnol. 2021 Apr 12.
      In contrast to single-cell approaches for measuring gene expression and DNA accessibility, single-cell methods for analyzing histone modifications are limited by low sensitivity and throughput. Here, we combine the CUT&Tag technology, developed to measure bulk histone modifications, with droplet-based single-cell library preparation to produce high-quality single-cell data on chromatin modifications. We apply single-cell CUT&Tag (scCUT&Tag) to tens of thousands of cells of the mouse central nervous system and probe histone modifications characteristic of active promoters, enhancers and gene bodies (H3K4me3, H3K27ac and H3K36me3) and inactive regions (H3K27me3). These scCUT&Tag profiles were sufficient to determine cell identity and deconvolute regulatory principles such as promoter bivalency, spreading of H3K4me3 and promoter-enhancer connectivity. We also used scCUT&Tag to investigate the single-cell chromatin occupancy of transcription factor OLIG2 and the cohesin complex component RAD21. Our results indicate that analysis of histone modifications and transcription factor occupancy at single-cell resolution provides unique insights into epigenomic landscapes in the central nervous system.
    DOI:  https://doi.org/10.1038/s41587-021-00869-9
  10. Cell. 2021 Apr 07. pii: S0092-8674(21)00353-6. [Epub ahead of print]
      A general approach for heritably altering gene expression has the potential to enable many discovery and therapeutic efforts. Here, we present CRISPRoff-a programmable epigenetic memory writer consisting of a single dead Cas9 fusion protein that establishes DNA methylation and repressive histone modifications. Transient CRISPRoff expression initiates highly specific DNA methylation and gene repression that is maintained through cell division and differentiation of stem cells to neurons. Pairing CRISPRoff with genome-wide screens and analysis of chromatin marks establishes rules for heritable gene silencing. We identify single guide RNAs (sgRNAs) capable of silencing the large majority of genes including those lacking canonical CpG islands (CGIs) and reveal a wide targeting window extending beyond annotated CGIs. The broad ability of CRISPRoff to initiate heritable gene silencing even outside of CGIs expands the canonical model of methylation-based silencing and enables diverse applications including genome-wide screens, multiplexed cell engineering, enhancer silencing, and mechanistic exploration of epigenetic inheritance.
    Keywords:  CRISPR; DNA methylation; cell therapy; dCas9; epigenetics
    DOI:  https://doi.org/10.1016/j.cell.2021.03.025
  11. Oncogene. 2021 Apr 12.
      The molecular mechanisms of luminal cell differentiation are not understood well enough to determine how differentiation goes awry during oncogenesis. Using RNA-Seq analysis, we discovered that CREB1 plays a central role in maintaining new luminal cell survival and that oncogenesis dramatically changes the CREB1-induced transcriptome. CREB1 is active in luminal cells, but not basal cells. We identified ING4 and its E3 ligase, JFK, as CREB1 transcriptional targets in luminal cells. During luminal cell differentiation, transient induction of ING4 expression is followed by a peak in CREB1 activity, while JFK increases concomitantly with CREB1 activation. Transient expression of ING4 is required for luminal cell induction; however, failure to properly down-regulate ING4 leads to luminal cell death. Consequently, blocking CREB1 increased ING4 expression, suppressed JFK, and led to luminal cell death. Thus, CREB1 is responsible for the suppression of ING4 required for luminal cell survival and maintenance. Oncogenic transformation by suppressing PTEN resulted in constitutive activation of CREB1. However, the tumor cells could no longer fully differentiate into luminal cells, failed to express ING4, and displayed a unique CREB1 transcriptome. Blocking CREB1 in tumorigenic cells suppressed tumor growth in vivo, rescued ING4 expression, and restored luminal cell formation, but ultimately induced luminal cell death. IHC of primary prostate tumors demonstrated a strong correlation between loss of ING4 and loss of PTEN. This is the first study to define a molecular mechanism whereby oncogenic loss of PTEN, leading to aberrant CREB1 activation, suppresses ING4 expression causing disruption of luminal cell differentiation.
    DOI:  https://doi.org/10.1038/s41388-021-01772-y
  12. PLoS Genet. 2021 Apr 12. 17(4): e1009498
      In mammals, cellular identity is defined through strict regulation of chromatin modifications and DNA methylation that control gene expression. Methylation of cytosines at CpG sites in the genome is mainly associated with suppression; however, the reason for enhancer-specific methylation is not fully understood. We used sequential ChIP-bisulfite-sequencing for H13K4me1 and H3K27ac histone marks. By collecting data from the same genomic region, we identified enhancers differentially methylated between these two marks. We observed a global gain of CpG methylation primarily in H3K4me1-marked nucleosomes during mouse embryonic stem cell differentiation. This gain occurred largely in enhancer regions that regulate genes critical for differentiation. The higher levels of DNA methylation in H3K4me1- versus H3K27ac-marked enhancers, despite it being the same genomic region, indicates cellular heterogeneity of enhancer states. Analysis of single-cell RNA-seq profiles demonstrated that this heterogeneity correlates with gene expression during differentiation. Furthermore, heterogeneity of enhancer methylation correlates with transcription start site methylation. Our results provide insights into enhancer-based functional variation in complex biological systems.
    DOI:  https://doi.org/10.1371/journal.pgen.1009498
  13. Nat Genet. 2021 Apr 15.
      Known fetal hemoglobin (HbF) silencers have potential on-target liabilities for rational β-hemoglobinopathy therapeutic inhibition. Here, through transcription factor (TF) CRISPR screening, we identify zinc-finger protein (ZNF) 410 as an HbF repressor. ZNF410 does not bind directly to the genes encoding γ-globins, but rather its chromatin occupancy is concentrated solely at CHD4, encoding the NuRD nucleosome remodeler, which is itself required for HbF repression. CHD4 has two ZNF410-bound regulatory elements with 27 combined ZNF410 binding motifs constituting unparalleled genomic clusters. These elements completely account for the effects of ZNF410 on fetal globin repression. Knockout of ZNF410 or its mouse homolog Zfp410 reduces CHD4 levels by 60%, enough to substantially de-repress HbF while eluding cellular or organismal toxicity. These studies suggest a potential target for HbF induction for β-hemoglobin disorders with a wide therapeutic index. More broadly, ZNF410 represents a special class of gene regulator, a conserved TF with singular devotion to regulation of a chromatin subcomplex.
    DOI:  https://doi.org/10.1038/s41588-021-00843-w
  14. Nature. 2021 Apr 14.
      Fundamental features of 3D genome organization are established de novo in the early embryo, including clustering of pericentromeric regions, the folding of chromosome arms and the segregation of chromosomes into active (A-) and inactive (B-) compartments. However, the molecular mechanisms that drive de novo organization remain unknown1,2. Here, by combining chromosome conformation capture (Hi-C), chromatin immunoprecipitation with high-throughput sequencing (ChIP-seq), 3D DNA fluorescence in situ hybridization (3D DNA FISH) and polymer simulations, we show that heterochromatin protein 1a (HP1a) is essential for de novo 3D genome organization during Drosophila early development. The binding of HP1a at pericentromeric heterochromatin is required to establish clustering of pericentromeric regions. Moreover, HP1a binding within chromosome arms is responsible for overall chromosome folding and has an important role in the formation of B-compartment regions. However, depletion of HP1a does not affect the A-compartment, which suggests that a different molecular mechanism segregates active chromosome regions. Our work identifies HP1a as an epigenetic regulator that is involved in establishing the global structure of the genome in the early embryo.
    DOI:  https://doi.org/10.1038/s41586-021-03460-z
  15. Nat Commun. 2021 04 13. 12(1): 2220
      The acidic activation domain (AD) of yeast transcription factor Gal4 plays a dual role in transcription repression and activation through binding to Gal80 repressor and Mediator subunit Med15. The activation function of Gal4 arises from two hydrophobic regions within the 40-residue AD. We show by NMR that each AD region binds the Mediator subunit Med15 using a "fuzzy" protein interface. Remarkably, comparison of chemical shift perturbations shows that Gal4 and Gcn4, two intrinsically disordered ADs of different sequence, interact nearly identically with Med15. The finding that two ADs of different sequence use an identical fuzzy binding mechanism shows a common sequence-independent mechanism for AD-Mediator binding, similar to interactions within a hydrophobic cloud. In contrast, the same region of Gal4 AD interacts strongly with Gal80 via a distinct structured complex, implying that the structured binding partner of an intrinsically disordered protein dictates the type of protein-protein interaction.
    DOI:  https://doi.org/10.1038/s41467-021-22441-4
  16. BMC Bioinformatics. 2021 Apr 15. 22(1): 193
      BACKGROUND: ChIP-seq combines chromatin immunoprecipitation assays with sequencing and identifies genome-wide binding sites for DNA binding proteins. While many binding sites have strong ChIP-seq 'peak' observations and are well captured, there are still regions bound by proteins weakly, with a relatively low ChIP-seq signal enrichment. These weak binding sites, especially those at promoters and enhancers, are functionally important because they also regulate nearby gene expression. Yet, it remains a challenge to accurately identify weak binding sites in ChIP-seq data due to the ambiguity in differentiating these weak binding sites from the amplified background DNAs.RESULTS: ChIP-BIT2 ( http://sourceforge.net/projects/chipbitc/ ) is a software package for ChIP-seq peak detection. ChIP-BIT2 employs a mixture model integrating protein and control ChIP-seq data and predicts strong or weak protein binding sites at promoters, enhancers, or other genomic locations. For binding sites at gene promoters, ChIP-BIT2 simultaneously predicts their target genes. ChIP-BIT2 has been validated on benchmark regions and tested using large-scale ENCODE ChIP-seq data, demonstrating its high accuracy and wide applicability.
    CONCLUSION: ChIP-BIT2 is an efficient ChIP-seq peak caller. It provides a better lens to examine weak binding sites and can refine or extend the existing binding site collection, providing additional regulatory regions for decoding the mechanism of gene expression regulation.
    DOI:  https://doi.org/10.1186/s12859-021-04108-5
  17. Cell Rep. 2021 Apr 13. pii: S2211-1247(21)00291-6. [Epub ahead of print]35(2): 108977
      Accumulation of topological stress in the form of DNA supercoiling is inherent to the advance of RNA polymerase II (Pol II) and needs to be resolved by DNA topoisomerases to sustain productive transcriptional elongation. Topoisomerases are therefore considered positive facilitators of transcription. Here, we show that, in contrast to this general assumption, human topoisomerase IIα (TOP2A) activity at promoters represses transcription of immediate early genes such as c-FOS, maintaining them under basal repressed conditions. Thus, TOP2A inhibition creates a particular topological context that results in rapid release from promoter-proximal pausing and transcriptional upregulation, which mimics the typical bursting behavior of these genes in response to physiological stimulus. We therefore describe the control of promoter-proximal pausing by TOP2A as a layer for the regulation of gene expression, which can act as a molecular switch to rapidly activate transcription, possibly by regulating the accumulation of DNA supercoiling at promoter regions.
    Keywords:  DNA supercoiling; DNA topoisomerases; DNA topology; gene expression; promoter-proximal pausing; transcription elongation
    DOI:  https://doi.org/10.1016/j.celrep.2021.108977
  18. Gut. 2021 Apr 12. pii: gutjnl-2020-321397. [Epub ahead of print]
      OBJECTIVE: Molecular taxonomy of tumours is the foundation of personalised medicine and is becoming of paramount importance for therapeutic purposes. Four transcriptomics-based classification systems of pancreatic ductal adenocarcinoma (PDAC) exist, which consistently identified a subtype of highly aggressive PDACs with basal-like features, including ΔNp63 expression and loss of the epithelial master regulator GATA6. We investigated the precise molecular events driving PDAC progression and the emergence of the basal programme.DESIGN: We combined the analysis of patient-derived transcriptomics datasets and tissue samples with mechanistic experiments using a novel dual-recombinase mouse model for Gata6 deletion at late stages of KRasG12D-driven pancreatic tumorigenesis (Gata6LateKO).
    RESULTS: This comprehensive human-to-mouse approach showed that GATA6 loss is necessary, but not sufficient, for the expression of ΔNp63 and the basal programme in patients and in mice. The concomitant loss of HNF1A and HNF4A, likely through epigenetic silencing, is required for the full phenotype switch. Moreover, Gata6 deletion in mice dramatically increased the metastatic rate, with a propensity for lung metastases. Through RNA-Seq analysis of primary cells isolated from mouse tumours, we show that Gata6 inhibits tumour cell plasticity and immune evasion, consistent with patient-derived data, suggesting that GATA6 works as a barrier for acquiring the fully developed basal and metastatic phenotype.
    CONCLUSIONS: Our work provides both a mechanistic molecular link between the basal phenotype and metastasis and a valuable preclinical tool to investigate the most aggressive subtype of PDAC. These data, therefore, are important for understanding the pathobiological features underlying the heterogeneity of pancreatic cancer in both mice and human.
    Keywords:  epithelial differentiation; molecular mechanisms; pancreatic cancer
    DOI:  https://doi.org/10.1136/gutjnl-2020-321397
  19. Nat Commun. 2021 Apr 16. 12(1): 2298
      Neutrophils play fundamental roles in innate immune response, shape adaptive immunity, and are a potentially causal cell type underpinning genetic associations with immune system traits and diseases. Here, we profile the binding of myeloid master regulator PU.1 in primary neutrophils across nearly a hundred volunteers. We show that variants associated with differential PU.1 binding underlie genetically-driven differences in cell count and susceptibility to autoimmune and inflammatory diseases. We integrate these results with other multi-individual genomic readouts, revealing coordinated effects of PU.1 binding variants on the local chromatin state, enhancer-promoter contacts and downstream gene expression, and providing a functional interpretation for 27 genes underlying immune traits. Collectively, these results demonstrate the functional role of PU.1 and its target enhancers in neutrophil transcriptional control and immune disease susceptibility.
    DOI:  https://doi.org/10.1038/s41467-021-22548-8
  20. Cell Rep. 2021 Apr 13. pii: S2211-1247(21)00295-3. [Epub ahead of print]35(2): 108981
      Despite the central role of chromosomal context in gene transcription, human noncoding DNA variants are generally studied outside of their genomic location. This limits our understanding of disease-causing regulatory variants. INS promoter mutations cause recessive neonatal diabetes. We show that all INS promoter point mutations in 60 patients disrupt a CC dinucleotide, whereas none affect other elements important for episomal promoter function. To model CC mutations, we humanized an ∼3.1-kb region of the mouse Ins2 gene. This recapitulated developmental chromatin states and cell-specific transcription. A CC mutant allele, however, abrogated active chromatin formation during pancreas development. A search for transcription factors acting through this element revealed that another neonatal diabetes gene product, GLIS3, has a pioneer-like ability to derepress INS chromatin, which is hampered by the CC mutation. Our in vivo analysis, therefore, connects two human genetic defects in an essential mechanism for developmental activation of the INS gene.
    Keywords:  GLIS3; HIP; INS promoter; mouse model; neonatal diabetes; regulatory element
    DOI:  https://doi.org/10.1016/j.celrep.2021.108981
  21. NAR Genom Bioinform. 2021 Jun;3(2): lqab022
      Identifying active transcriptional regulators (TRs) associating with cis-regulatory elements in the genome to regulate gene expression is a key task in gene regulation research. TR binding profiles from numerous public ChIP-seq data can be utilized for association analysis with query data for TR identification, as an alternative to DNA sequence motif analysis. However, integration of the massive ChIP-seq datasets has been a major challenge in such approaches. Here we present BARTweb, an interactive web server for identifying TRs whose genomic binding patterns associate with input genomic features, by leveraging over 13 000 public ChIP-seq datasets for human and mouse. Using an updated binding analysis for regulation of transcription (BART) algorithm, BARTweb can identify functional TRs that regulate a gene set, have a binding profile correlated with a ChIP-seq profile or are enriched in a genomic region set, without a priori information of the cell type. BARTweb can be a useful web server for performing functional analysis of gene regulation. BARTweb is freely available at http://bartweb.org and the source code is available at https://github.com/zanglab/bart2.
    DOI:  https://doi.org/10.1093/nargab/lqab022
  22. Nat Commun. 2021 Apr 16. 12(1): 2282
      Acheiropodia, congenital limb truncation, is associated with homozygous deletions in the LMBR1 gene around ZRS, an enhancer regulating SHH during limb development. How these deletions lead to this phenotype is unknown. Using whole-genome sequencing, we fine-mapped the acheiropodia-associated region to 12 kb and show that it does not function as an enhancer. CTCF and RAD21 ChIP-seq together with 4C-seq and DNA FISH identify three CTCF sites within the acheiropodia-deleted region that mediate the interaction between the ZRS and the SHH promoter. This interaction is substituted with other CTCF sites centromeric to the ZRS in the disease state. Mouse knockouts of the orthologous 12 kb sequence have no apparent abnormalities, showcasing the challenges in modelling CTCF alterations in animal models due to inherent motif differences between species. Our results show that alterations in CTCF motifs can lead to a Mendelian condition due to altered enhancer-promoter interactions.
    DOI:  https://doi.org/10.1038/s41467-021-22470-z
  23. Nucleic Acids Res. 2021 Apr 15. pii: gkab246. [Epub ahead of print]
      Hexasomes and tetrasomes are intermediates in nucleosome assembly and disassembly. Their formation is promoted by histone chaperones, ATP-dependent remodelers, and RNA polymerase II. In addition, hexasomes are maintained in transcribed genes and could be an important regulatory factor. While nucleosome composition has been shown to affect the structure and accessibility of DNA, its influence on histone tails is largely unknown. Here, we investigate the conformational dynamics of the H3 tail in the hexasome and tetrasome. Using a combination of NMR spectroscopy, MD simulations, and trypsin proteolysis, we find that the conformational ensemble of the H3 tail is regulated by nucleosome composition. As has been found for the nucleosome, the H3 tails bind robustly to DNA within the hexasome and tetrasome, but upon loss of the H2A/H2B dimer, we determined that the adjacent H3 tail has an altered conformational ensemble, increase in dynamics, and increase in accessibility. Similar to observations of DNA dynamics, this is seen to be asymmetric in the hexasome. Our results indicate that nucleosome composition has the potential to regulate chromatin signaling and ultimately help shape the chromatin landscape.
    DOI:  https://doi.org/10.1093/nar/gkab246
  24. Nat Commun. 2021 04 14. 12(1): 2242
      Breast cancer is a heterogeneous disease, affecting over 3.5 million women worldwide, yet the functional role of cis-regulatory elements including super-enhancers in different breast cancer subtypes remains poorly characterized. Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with a poor prognosis. Here we apply integrated epigenomic and transcriptomic profiling to uncover super-enhancer heterogeneity between breast cancer subtypes, and provide clinically relevant biological insights towards TNBC. Using CRISPR/Cas9-mediated gene editing, we identify genes that are specifically regulated by TNBC-specific super-enhancers, including FOXC1 and MET, thereby unveiling a mechanism for specific overexpression of the key oncogenes in TNBC. We also identify ANLN as a TNBC-specific gene regulated by super-enhancer. Our studies reveal a TNBC-specific epigenomic landscape, contributing to the dysregulated oncogene expression in breast tumorigenesis.
    DOI:  https://doi.org/10.1038/s41467-021-22445-0
  25. BMC Biol. 2021 Apr 16. 19(1): 79
      BACKGROUND: Rheumatoid arthritis (RA) is a chronic, systemic autoimmune disease that involves a variety of cell types. However, how the epigenetic dysregulations of peripheral immune cells contribute to the pathogenesis of RA still remains largely unclear.RESULTS: Here, we analysed the genome-wide active DNA regulatory elements of four major immune cells, namely monocytes, B cells, CD4+ T cells and CD8+ T cells, in peripheral blood of RA patients, osteoarthritis (OA) patients and healthy donors using Assay of Transposase Accessible Chromatin with sequencing (ATAC-seq). We found a strong RA-associated chromatin dysregulation signature in monocytes, but no other examined cell types. Moreover, we found that serum C-reactive protein (CRP) can induce the RA-associated chromatin dysregulation in monocytes via in vitro experiments. And the extent of this dysregulation was regulated through the transcription factor FRA2.
    CONCLUSIONS: Together, our study revealed a CRP-induced pathogenic chromatin dysregulation signature in monocytes from RA patients and predicted the responsible signalling pathway as potential therapeutic targets for the disease.
    Keywords:  C-reactive protein; Chromatin dysregulation; FRA2; Monocytes; Rheumatoid arthritis (RA)
    DOI:  https://doi.org/10.1186/s12915-021-01011-6
  26. Elife. 2021 04 13. pii: e62293. [Epub ahead of print]10
      Aging is associated with complex molecular and cellular processes that are poorly understood. Here we leveraged the Tabula Muris Senis single-cell RNA-seq data set to systematically characterize gene expression changes during aging across diverse cell types in the mouse. We identified aging-dependent genes in 76 tissue-cell types from 23 tissues and characterized both shared and tissue-cell-specific aging behaviors. We found that the aging-related genes shared by multiple tissue-cell types also change their expression congruently in the same direction during aging in most tissue-cell types, suggesting a coordinated global aging behavior at the organismal level. Scoring cells based on these shared aging genes allowed us to contrast the aging status of different tissues and cell types from a transcriptomic perspective. In addition, we identified genes that exhibit age-related expression changes specific to each functional category of tissue-cell types. Altogether, our analyses provide one of the most comprehensive and systematic characterizations of the molecular signatures of aging across diverse tissue-cell types in a mammalian system.
    Keywords:  aging; computation; computational biology; mouse; single cell; systems biology
    DOI:  https://doi.org/10.7554/eLife.62293
  27. Genome Res. 2021 Apr 14. pii: gr.260893.120. [Epub ahead of print]
      Recently, multiple single-cell assays were developed for detecting histone marks at the single-cell levels. These techniques are either limited by the low cell throughput or sparse reads which limit their applications. To address these problems, we introduce indexing single-cell immunocleavage sequencing (iscChIC-seq), a multiplex indexing method based on TdT terminal transferase and T4 DNA ligase mediated barcoding strategy and single-cell ChIC-seq, which is capable of readily analyzing histone modifications across tens of thousands of single cells in one experiment. Application of iscChIC-seq to profiling H3K4me3 and H3K27me3 in human white blood cells (WBCs) enabled successful detection of more than 10,000 single cells for each histone modification with 11K and 45K nonredundant reads per cell, respectively. Cluster analysis of these data allowed identification of monocytes, T cells, B cells, and NK cells from WBCs. The cell types annotated from H3K4me3 single-cell data are specifically correlated with the cell types annotated from H3K27me3 single-cell data. Our data indicate that iscChIC-seq is a reliable technique for profiling histone modifications in a large number of single cells, which may find broad applications in studying cellular heterogeneity and differentiation status in complex developmental and disease systems.
    DOI:  https://doi.org/10.1101/gr.260893.120
  28. PLoS One. 2021 ;16(4): e0249985
      Understanding the factors that underlie the epigenetic regulation of genes is crucial to understand the gene regulatory machinery as a whole. Several experimental and computational studies examined the relationship between different factors involved. Here we investigate the relationship between transcription factors (TFs) and histone modifications (HMs), based on ChIP-seq data in cell lines. As it was shown that gene regulation by TFs differs depending on the CpG class of a promoter, we study the impact of the CpG content in promoters on the associations between TFs and HMs. We suggest an approach based on sparse linear regression models to infer associations between TFs and HMs with respect to CpG content. A study of the partial correlation of HMs for the two classes of high and low CpG content reveals possible CpG dependence and potential candidates for confounding factors in our models. We show that the models are accurate, inferred associations reflect known biological relationships, and we give new insight into associations with respect to CpG content. Moreover, analysis of a ChIP-seq dataset in HepG2 cells of the HM H3K122ac, an HM about little is known, reveals novel TF associations and supports a previously established link to active transcription.
    DOI:  https://doi.org/10.1371/journal.pone.0249985
  29. Oncogene. 2021 Apr 16.
      Colorectal cancer (CRC) is one of the leading cancers worldwide, accounting for high morbidity and mortality. The mechanisms governing tumor growth and metastasis in CRC require detailed investigation. The results of the present study indicated that the transcription factor (TF) myocyte enhancer factor 2A (MEF2A) plays a dual role in promoting proliferation and metastasis of CRC by inducing the epithelial-mesenchymal transition (EMT) and activation of WNT/β-catenin signaling. Aberrant expression of MEF2A in CRC clinical specimens was significantly associated with poor prognosis and metastasis. Functionally, MEF2A directly binds to the promoter region to initiate the transcription of ZEB2 and CTNNB1. Simultaneous activation of the expression of EMT-related TFs and Wnt/β-catenin signaling by MEF2A overexpression induced the EMT and increased the frequency of tumor formation and metastasis. The present study identified a new critical oncogene involved in the growth and metastasis of CRC, providing a potential novel therapeutic target for CRC intervention.
    DOI:  https://doi.org/10.1038/s41388-021-01774-w
  30. Nat Commun. 2021 04 12. 12(1): 2151
      The rapid increase in the amount of genomic data provides researchers with an opportunity to integrate diverse datasets and annotations when addressing a wide range of biological questions. However, genomic datasets are deposited on different platforms and are stored in numerous formats from multiple genome builds, which complicates the task of collecting, annotating, transforming, and integrating data as needed. Here, we developed Go Get Data (GGD) as a fast, reproducible approach to installing standardized data recipes. GGD is available on Github ( https://gogetdata.github.io/ ), is extendable to other data types, and can streamline the complexities typically associated with data integration, saving researchers time and improving research reproducibility.
    DOI:  https://doi.org/10.1038/s41467-021-22381-z
  31. Proc Natl Acad Sci U S A. 2021 Apr 20. pii: e2017148118. [Epub ahead of print]118(16):
      Sex can be an important determinant of cancer phenotype, and exploring sex-biased tumor biology holds promise for identifying novel therapeutic targets and new approaches to cancer treatment. In an established isogenic murine model of glioblastoma (GBM), we discovered correlated transcriptome-wide sex differences in gene expression, H3K27ac marks, large Brd4-bound enhancer usage, and Brd4 localization to Myc and p53 genomic binding sites. These sex-biased gene expression patterns were also evident in human glioblastoma stem cells (GSCs). These observations led us to hypothesize that Brd4-bound enhancers might underlie sex differences in stem cell function and tumorigenicity in GBM. We found that male and female GBM cells exhibited sex-specific responses to pharmacological or genetic inhibition of Brd4. Brd4 knockdown or pharmacologic inhibition decreased male GBM cell clonogenicity and in vivo tumorigenesis while increasing both in female GBM cells. These results were validated in male and female patient-derived GBM cell lines. Furthermore, analysis of the Cancer Therapeutic Response Portal of human GBM samples segregated by sex revealed that male GBM cells are significantly more sensitive to BET (bromodomain and extraterminal) inhibitors than are female cells. Thus, Brd4 activity is revealed to drive sex differences in stem cell and tumorigenic phenotypes, which can be abrogated by sex-specific responses to BET inhibition. This has important implications for the clinical evaluation and use of BET inhibitors.
    Keywords:  BET inhibitors; Brd4-bound enhancers; glioblastoma; sex differences; sex-specific transcriptional programs
    DOI:  https://doi.org/10.1073/pnas.2017148118