bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2022–08–14
23 papers selected by
Connor Rogerson, University of Cambridge
  1. HIC2 controls developmental hemoglobin switching by repressing BCL11A transcription.
  2. TET enzymes regulate skeletal development through increasing chromatin accessibility of RUNX2 target genes.
  3. Temporal analysis of enhancers during mouse cerebellar development reveals dynamic and novel regulatory functions.
  4. Single-cell diploid Hi-C reveals the role of spatial aggregations in complex rearrangements and KMT2A fusions in leukemia.
  5. scBasset: sequence-based modeling of single-cell ATAC-seq using convolutional neural networks.
  6. Concentration-dependent transcriptional switching through a collective action of cis-elements.
  7. Epigenetic reader SP140 loss of function drives Crohn's disease due to uncontrolled macrophage topoisomerases.
  8. Nested epistasis enhancer networks for robust genome regulation.
  9. Single base-pair resolution analysis of DNA binding motif with MoMotif reveals an oncogenic function of CTCF zinc-finger 1 mutation.
  10. SHP-1 dephosphorylates histone H2B to facilitate its ubiquitination during transcription.
  11. A Distinct Chromatin State Drives Therapeutic Resistance in Invasive Lobular Breast Cancer.
  12. The SWI/SNF chromatin remodeling factor DPF3 regulates metastasis of ccRCC by modulating TGF-β signaling.
  13. PRC1-mediated epigenetic programming is required to generate the ovarian reserve.
  14. Transcription-coupled and epigenome-encoded mechanisms direct H3K4 methylation.
  15. Endogenous Retroviral Sequences Behave as Putative Enhancers Controlling Gene Expression through HP1-Regulated Long-Range Chromatin Interactions.
  16. HaloChIP-seq for Antibody-Independent Mapping of Mouse Transcription Factor Cistromes in vivo.
  17. Adrenergic receptor signaling induced by Klf15, a regulator of regeneration enhancer, promotes kidney reconstruction.
  18. Chromatin accessibility of primary human cancers ties regional mutational processes and signatures with tissues of origin.
  19. GREAP: a comprehensive enrichment analysis software for human genomic regions.
  20. Generation of functional hepatocytes by forward programming with nuclear receptors.
  21. Transcription factors perform a 2-step search of the nucleus.
  22. Differential dependencies of human RNA polymerase II promoters on TBP, TAF1, TFIIB and XPB.
  23. FTO promotes clear cell renal cell carcinoma progression via upregulation of PDK1 through an m6A dependent pathway.
  1. Nat Genet. 2022 Aug 08.
    HIC2 controls developmental hemoglobin switching by repressing BCL11A transcription.
    Peng Huang, Scott A Peslak, Ren Ren, Eugene Khandros, Kunhua Qin, Cheryl A Keller, Belinda Giardine, Henry W Bell, Xianjiang Lan, Malini Sharma, John R Horton, Osheiza Abdulmalik, Stella T Chou, Junwei Shi, Merlin Crossley, Ross C Hardison, Xiaodong Cheng, Gerd A Blobel.
      The fetal-to-adult switch in hemoglobin production is a model of developmental gene control with relevance to the treatment of hemoglobinopathies. The expression of transcription factor BCL11A, which represses fetal β-type globin (HBG) genes in adult erythroid cells, is predominantly controlled at the transcriptional level but the underlying mechanism is unclear. We identify HIC2 as a repressor of BCL11A transcription. HIC2 and BCL11A are reciprocally expressed during development. Forced expression of HIC2 in adult erythroid cells inhibits BCL11A transcription and induces HBG expression. HIC2 binds to erythroid BCL11A enhancers to reduce chromatin accessibility and binding of transcription factor GATA1, diminishing enhancer activity and enhancer-promoter contacts. DNA-binding and crystallography studies reveal direct steric hindrance as one mechanism by which HIC2 inhibits GATA1 binding at a critical BCL11A enhancer. Conversely, loss of HIC2 in fetal erythroblasts increases enhancer accessibility, GATA1 binding and BCL11A transcription. HIC2 emerges as an evolutionarily conserved regulator of hemoglobin switching via developmental control of BCL11A.
    DOI:  https://doi.org/10.1038/s41588-022-01152-6
  2. Nat Commun. 2022 Aug 11. 13(1): 4709
    TET enzymes regulate skeletal development through increasing chromatin accessibility of RUNX2 target genes.
    Lijun Wang, Xiuling You, Dengfeng Ruan, Rui Shao, Hai-Qiang Dai, Weiliang Shen, Guo-Liang Xu, Wanlu Liu, Weiguo Zou.
      The Ten-eleven translocation (TET) family of dioxygenases mediate cytosine demethylation by catalyzing the oxidation of 5-methylcytosine (5mC). TET-mediated DNA demethylation controls the proper differentiation of embryonic stem cells and TET members display functional redundancy during early gastrulation. However, it is unclear if TET proteins have functional significance in mammalian skeletal development. Here, we report that Tet genes deficiency in mesoderm mesenchymal stem cells results in severe defects of bone development. The existence of any single Tet gene allele can support early bone formation, suggesting a functional redundancy of TET proteins. Integrative analyses of RNA-seq, Whole Genome Bisulfite Sequencing (WGBS), 5hmC-Seal and Assay for Transposase-Accessible Chromatin (ATAC-seq) demonstrate that TET-mediated demethylation increases the chromatin accessibility of target genes by RUNX2 and facilities RUNX2-regulated transcription. In addition, TET proteins interact with RUNX2 through their catalytic domain to regulate cytosine methylation around RUNX2 binding region. The catalytic domain is indispensable for TET enzymes to regulate RUNX2 transcription activity on its target genes and to regulate bone development. These results demonstrate that TET enzymes function to regulate RUNX2 activity and maintain skeletal homeostasis.
    DOI:  https://doi.org/10.1038/s41467-022-32138-x
  3. Elife. 2022 Aug 09. pii: e74207. [Epub ahead of print]11
    Temporal analysis of enhancers during mouse cerebellar development reveals dynamic and novel regulatory functions.
    FANTOM 5 Consortium
      We have identified active enhancers in the mouse cerebellum at embryonic and postnatal stages which provides a view of novel enhancers active during cerebellar development. The majority of cerebellar enhancers have dynamic activity between embryonic and postnatal development. Cerebellar enhancers were enriched for neural transcription factor binding sites with temporally specific expression. Putative gene targets displayed spatially restricted expression patterns, indicating cell-type specific expression regulation. Functional analysis of target genes indicated that enhancers regulate processes spanning several developmental epochs such as specification, differentiation and maturation. We use these analyses to discover one novel regulator and one novel marker of cerebellar development: Bhlhe22 and Pax3, respectively. We identified an enrichment of de novo mutations and variants associated with autism spectrum disorder in cerebellar enhancers. Furthermore, by comparing our data with relevant brain development ENCODE histone profiles and cerebellar single-cell datasets we have been able to generalize and expand on the presented analyses, respectively. We have made the results of our analyses available online in the Developing Mouse Cerebellum Enhancer Atlas (https://goldowitzlab.shinyapps.io/developing_mouse_cerebellum_enhancer_atlas/), where our dataset can be efficiently queried, curated and exported by the scientific community to facilitate future research efforts. Our study provides a valuable resource for studying the dynamics of gene expression regulation by enhancers in the developing cerebellum and delivers a rich dataset of novel gene-enhancer associations providing a basis for future in-depth studies in the cerebellum.
    Keywords:  developmental biology; mouse; neuroscience
    DOI:  https://doi.org/10.7554/eLife.74207
  4. Genome Biol. 2022 Aug 09. 23(1): 173
    Single-cell diploid Hi-C reveals the role of spatial aggregations in complex rearrangements and KMT2A fusions in leukemia.
    Zhihao Xing, Huirong Mai, Xiaorong Liu, Xiaoying Fu, Xingliang Zhang, Lichun Xie, Yunsheng Chen, Adam Shlien, Feiqiu Wen.
       BACKGROUND: Simple translocations and complex rearrangements are formed through illegitimate ligations of double-strand breaks of fusion partners and lead to generation of oncogenic fusion genes that affect cellular function. The contact first hypothesis states that fusion partners tend to colocalize prior to fusion in normal cells. Here we test this hypothesis at the single-cell level and explore the underlying mechanism.
    RESULTS: By analyzing published single-cell diploid Hi-C datasets, we find partner genes fused in leukemia exhibit smaller spatial distances than those fused in solid tumor and control gene pairs. Intriguingly, multiple partners tend to colocalize with KMT2A in the same cell. 3D genome architecture has little association with lineage decision of KMT2A fusion types in leukemia. Besides simple translocations, complex rearrangement-related KMT2A fusion genes (CRGs) also show closer proximity and belong to a genome-wide mutual proximity network. We find CRGs are co-expressed, co-localized, and enriched in the targets of the transcriptional factor RUNX1, suggesting they may be involved in RUNX1-mediated transcription factories. Knockdown of RUNX1 leads to significantly fewer contacts among CRGs. We also find CRGs are enriched in active transcriptional regions and loop anchors, and exhibit high levels of TOP2-mediated DNA breakages. Inhibition of transcription leads to reduced DNA breakages of CRGs.
    CONCLUSIONS: Our results demonstrate KMT2A partners and CRGs may form dynamic and multipartite spatial clusters in individual cells that may be involved in RUNX1-mediated transcription factories, wherein massive DNA damages and illegitimate ligations of genes may occur, leading to complex rearrangements and KMT2A fusions in leukemia.
    Keywords:  Complex rearrangements; Fusions; Genomic rearrangements; Leukemia; Single-cell Hi-C
    DOI:  https://doi.org/10.1186/s13059-022-02740-9
  5. Nat Methods. 2022 Aug 08.
    scBasset: sequence-based modeling of single-cell ATAC-seq using convolutional neural networks.
    Han Yuan, David R Kelley.
      Single-cell assay for transposase-accessible chromatin using sequencing (scATAC) shows great promise for studying cellular heterogeneity in epigenetic landscapes, but there remain important challenges in the analysis of scATAC data due to the inherent high dimensionality and sparsity. Here we introduce scBasset, a sequence-based convolutional neural network method to model scATAC data. We show that by leveraging the DNA sequence information underlying accessibility peaks and the expressiveness of a neural network model, scBasset achieves state-of-the-art performance across a variety of tasks on scATAC and single-cell multiome datasets, including cell clustering, scATAC profile denoising, data integration across assays and transcription factor activity inference.
    DOI:  https://doi.org/10.1038/s41592-022-01562-8
  6. Sci Adv. 2022 Aug 12. 8(32): eabo6157
    Concentration-dependent transcriptional switching through a collective action of cis-elements.
    Kevin Rodriguez, Albert Do, Betul Senay-Aras, Mariano Perales, Mark Alber, Weitao Chen, G Venugopala Reddy.
      Gene expression specificity of homeobox transcription factors has remained paradoxical. WUSCHEL activates and represses CLAVATA3 transcription at lower and higher concentrations, respectively. We use computational modeling and experimental analysis to investigate the properties of the cis-regulatory module. We find that intrinsically each cis-element can only activate CLAVATA3 at a higher WUSCHEL concentration. However, together, they repress CLAVATA3 at higher WUSCHEL and activate only at lower WUSCHEL, showing that the concentration-dependent interactions among cis-elements regulate both activation and repression. Biochemical experiments show that two adjacent functional cis-elements bind WUSCHEL with higher affinity and dimerize at relatively lower levels. Moreover, increasing the distance between cis-elements prolongs WUSCHEL monomer binding window, resulting in higher CLAVATA3 activation. Our work showing a constellation of optimally spaced cis-elements of defined affinities determining activation and repression thresholds in regulating CLAVATA3 transcription provides a previously unknown mechanism of cofactor-independent regulation of transcription factor binding in mediating gene expression specificity.
    DOI:  https://doi.org/10.1126/sciadv.abo6157
  7. Cell. 2022 Aug 10. pii: S0092-8674(22)00841-8. [Epub ahead of print]
    Epigenetic reader SP140 loss of function drives Crohn's disease due to uncontrolled macrophage topoisomerases.
    Hajera Amatullah, Isabella Fraschilla, Sreehaas Digumarthi, Julie Huang, Fatemeh Adiliaghdam, Gracia Bonilla, Lai Ping Wong, Marie-Eve Rivard, Claudine Beauchamp, Virginie Mercier, Philippe Goyette, Ruslan I Sadreyev, Robert M Anthony, John D Rioux, Kate L Jeffrey.
      How mis-regulated chromatin directly impacts human immune disorders is poorly understood. Speckled Protein 140 (SP140) is an immune-restricted PHD and bromodomain-containing epigenetic "reader," and SP140 loss-of-function mutations associate with Crohn's disease (CD), multiple sclerosis (MS), and chronic lymphocytic leukemia (CLL). However, the relevance of these mutations and mechanisms underlying SP140-driven pathogenicity remains unexplored. Using a global proteomic strategy, we identified SP140 as a repressor of topoisomerases (TOPs) that maintains heterochromatin and macrophage fate. In humans and mice, SP140 loss resulted in unleashed TOP activity, de-repression of developmentally silenced genes, and ultimately defective microbe-inducible macrophage transcriptional programs and bacterial killing that drive intestinal pathology. Pharmacological inhibition of TOP1/2 rescued these defects. Furthermore, exacerbated colitis was restored with TOP1/2 inhibitors in Sp140-/- mice, but not wild-type mice, in vivo. Collectively, we identify SP140 as a TOP repressor and reveal repurposing of TOP inhibition to reverse immune diseases driven by SP140 loss.
    Keywords:  Crohn’s disease; PHD; SP140; Speckled Protein; bromodomain; chromatin; epigenetic reader; inflammatory bowel disease; macrophages; topoisomerase
    DOI:  https://doi.org/10.1016/j.cell.2022.06.048
  8. Science. 2022 Aug 11. eabk3512
    Nested epistasis enhancer networks for robust genome regulation.
    Xueqiu Lin, Yanxia Liu, Shuai Liu, Xiang Zhu, Lingling Wu, Yanyu Zhu, Dehua Zhao, Xiaoshu Xu, Augustine Chemparathy, Haifeng Wang, Yaqiang Cao, Muneaki Nakamura, Jasprina N Noordermeer, Marie La Russa, Wing Hung Wong, Keji Zhao, Lei S Qi.
      Mammalian genomes possess multiple enhancers spanning an ultralong distance (>megabases) to modulate important genes, yet it is unclear how these enhancers coordinate to achieve this task. Here, we combine multiplexed CRISPRi screening with machine learning to define quantitative enhancer-enhancer interactions. We find that the ultralong distance enhancer network possesses a nested multi-layer architecture that confers functional robustness of gene expression. Experimental characterization reveals that enhancer epistasis is maintained by three-dimensional chromosomal interactions and BRD4 condensation. Machine learning prediction of synergistic enhancers provides an effective strategy to identify non-coding variant pairs associated with pathogenic genes in diseases beyond Genome-Wide Association Studies (GWAS) analysis. Our work unveils nested epistasis enhancer networks, which can better explain enhancer functions within cells and in diseases.
    DOI:  https://doi.org/10.1126/science.abk3512
  9. Nucleic Acids Res. 2022 Aug 10. pii: gkac658. [Epub ahead of print]
    Single base-pair resolution analysis of DNA binding motif with MoMotif reveals an oncogenic function of CTCF zinc-finger 1 mutation.
    Benjamin Lebeau, Kaiqiong Zhao, Maika Jangal, Tiejun Zhao, Maria Guerra, Celia M T Greenwood, Michael Witcher.
      Defining the impact of missense mutations on the recognition of DNA motifs is highly dependent on bioinformatic tools that define DNA binding elements. However, classical motif analysis tools remain limited in their capacity to identify subtle changes in complex binding motifs between distinct conditions. To overcome this limitation, we developed a new tool, MoMotif, that facilitates a sensitive identification, at the single base-pair resolution, of complex, or subtle, alterations to core binding motifs, discerned from ChIP-seq data. We employed MoMotif to define the previously uncharacterized recognition motif of CTCF zinc-finger 1 (ZF1), and to further define the impact of CTCF ZF1 mutation on its association with chromatin. Mutations of CTCF ZF1 are exclusive to breast cancer and are associated with metastasis and therapeutic resistance, but the underlying mechanisms are unclear. Using MoMotif, we identified an extension of the CTCF core binding motif, necessitating a functional ZF1 to bind appropriately. Using a combination of ChIP-Seq and RNA-Seq, we discover that the inability to bind this extended motif drives an altered transcriptional program associated with the oncogenic phenotypes observed clinically. Our study demonstrates that MoMotif is a powerful new tool for comparative ChIP-seq analysis and characterising DNA-protein contacts.
    DOI:  https://doi.org/10.1093/nar/gkac658
  10. EMBO J. 2022 Aug 08. e109720
    SHP-1 dephosphorylates histone H2B to facilitate its ubiquitination during transcription.
    Prajakta Tathe, K V S Rammohan Chowdary, Krushna Chandra Murmu, Punit Prasad, Subbareddy Maddika.
      Dynamic regulation of phosphorylation and dephosphorylation of histones is essential for eukaryotic transcription, but the enzymes engaged in histone dephosphorylation are not fully explored. Here, we show that the tyrosine phosphatase SHP-1 dephosphorylates histone H2B and plays a critical role during transition from the initiation to the elongation stage of transcription. Nuclear-localized SHP-1 is associated with the Paf1 complex at chromatin and dephosphorylates H2B at tyrosine 121. Moreover, knockout of SHP-1, or expression of a mutant mimicking constitutive phosphorylation of H2B Y121, leads to a reduction in genome-wide H2B ubiquitination, which subsequently causes defects in RNA polymerase II-dependent transcription. Mechanistically, we demonstrate that Y121 phosphorylation precludes H2B's interaction with the E2 enzyme, indicating that SHP-1-mediated dephosphorylation of this residue may be a prerequisite for efficient H2B ubiquitination. Functionally, we find that SHP-1-mediated H2B dephosphorylation contributes to maintaining basal autophagic flux in cells through the efficient transcription of autophagy and lysosomal genes. Collectively, our study reveals an important modification of histone H2B regulated by SHP-1 that has a role during eukaryotic transcription.
    Keywords:  H2B ubiquitination; Paf1 complex; SHP-1; histone H2B; transcription
    DOI:  https://doi.org/10.15252/embj.2021109720
  11. Cancer Res. 2022 Aug 11. pii: CAN-21-3186. [Epub ahead of print]
    A Distinct Chromatin State Drives Therapeutic Resistance in Invasive Lobular Breast Cancer.
    Agostina Nardone, Xintao Qiu, Sandor Spisak, Zsuzsanna Nagy, Ariel Feiglin, Avery Feit, Gabriela Cohen Feit, Yingtian Xie, Alba Font-Tello, Cristina Guarducci, Francisco Hermida-Prado, Sudeepa Syamala, Klothilda Lim, Miguel Munoz Gomez, Matthew Pun, MacIntosh Cornwell, Weihan Liu, Aysegul Ors, Hisham Mohammed, Paloma Cejas, Jane B Brock, Matthew L Freedman, Eric P Winer, Xiaoyong Fu, Rachel Schiff, Henry W Long, Otto Metzger Filho, Rinath Jeselsohn.
      Most invasive lobular breast cancers (ILC) are of the luminal A subtype and are strongly hormone receptor positive. Yet, ILC is relatively resistant to tamoxifen and associated with inferior long-term outcomes compared to invasive ductal cancers (IDC). In this study, we sought to gain mechanistic insights into these clinical findings that are not explained by the genetic landscape of ILC and to identify strategies to improve patient outcomes. A comprehensive analysis of the epigenome of ILC in pre-clinical models and clinical samples showed that, compared to IDC, ILC harbored a distinct chromatin state linked to gained recruitment of FOXA1, a lineage-defining pioneer transcription factor. This resulted in an ILC-unique FOXA1-estrogen receptor (ER) axis that promoted the transcription of genes associated with tumor progression and poor outcomes. The ILC-unique FOXA1-ER axis led to retained ER chromatin binding after tamoxifen treatment, which facilitated tamoxifen resistance while remaining strongly dependent on ER signaling. Mechanistically, gained FOXA1 binding was associated with the auto-induction of FOXA1 in ILC through an ILC-unique FOXA1 binding site. Targeted silencing of this regulatory site resulted in the disruption of the feed-forward loop and growth inhibition in ILC. In summary, ILC is characterized by a unique chromatin state and FOXA1-ER axis that is associated with tumor progression, offering a novel mechanism of tamoxifen resistance. These results underscore the importance of conducting clinical trials dedicated to patients with ILC in order to optimize treatments in this breast cancer subtype.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-21-3186
  12. Nat Commun. 2022 Aug 09. 13(1): 4680
    The SWI/SNF chromatin remodeling factor DPF3 regulates metastasis of ccRCC by modulating TGF-β signaling.
    Huanhuan Cui, Hongyang Yi, Hongyu Bao, Ying Tan, Chi Tian, Xinyao Shi, Diwen Gan, Bin Zhang, Weizheng Liang, Rui Chen, Qionghua Zhu, Liang Fang, Xin Gao, Hongda Huang, Ruijun Tian, Silke R Sperling, Yuhui Hu, Wei Chen.
      DPF3, a component of the SWI/SNF chromatin remodeling complex, has been associated with clear cell renal cell carcinoma (ccRCC) in a genome-wide association study. However, the functional role of DPF3 in ccRCC development and progression remains unknown. In this study, we demonstrate that DPF3a, the short isoform of DPF3, promotes kidney cancer cell migration both in vitro and in vivo, consistent with the clinical observation that DPF3a is significantly upregulated in ccRCC patients with metastases. Mechanistically, DPF3a specifically interacts with SNIP1, via which it forms a complex with SMAD4 and p300 histone acetyltransferase (HAT), the major transcriptional regulators of TGF-β signaling pathway. Moreover, the binding of DPF3a releases the repressive effect of SNIP1 on p300 HAT activity, leading to the increase in local histone acetylation and the activation of cell movement related genes. Overall, our findings reveal a metastasis-promoting function of DPF3, and further establish the link between SWI/SNF components and ccRCC.
    DOI:  https://doi.org/10.1038/s41467-022-32472-0
  13. Nat Commun. 2022 Aug 10. 13(1): 4510
    PRC1-mediated epigenetic programming is required to generate the ovarian reserve.
    Mengwen Hu, Yu-Han Yeh, Yasuhisa Munakata, Hironori Abe, Akihiko Sakashita, So Maezawa, Miguel Vidal, Haruhiko Koseki, Neil Hunter, Richard M Schultz, Satoshi H Namekawa.
      The ovarian reserve defines the female reproductive lifespan, which in humans spans decades due to robust maintenance of meiotic arrest in oocytes residing in primordial follicles. Epigenetic reprogramming, including DNA demethylation, accompanies meiotic entry, but the chromatin changes that underpin the generation and preservation of ovarian reserves are poorly defined. We report that the Polycomb Repressive Complex 1 (PRC1) establishes repressive chromatin states in perinatal mouse oocytes that directly suppress the gene expression program of meiotic prophase-I and thereby enable the transition to dictyate arrest. PRC1 dysfuction causes depletion of the ovarian reserve and leads to premature ovarian failure. Our study demonstrates a fundamental role for PRC1-mediated gene silencing in female reproductive lifespan, and reveals a critical window of epigenetic programming required to establish ovarian reserve.
    DOI:  https://doi.org/10.1038/s41467-022-31759-6
  14. Nat Commun. 2022 Aug 11. 13(1): 4521
    Transcription-coupled and epigenome-encoded mechanisms direct H3K4 methylation.
    Satoyo Oya, Mayumi Takahashi, Kazuya Takashima, Tetsuji Kakutani, Soichi Inagaki.
      Mono-, di-, and trimethylation of histone H3 lysine 4 (H3K4me1/2/3) are associated with transcription, yet it remains controversial whether H3K4me1/2/3 promote or result from transcription. Our previous characterizations of Arabidopsis H3K4 demethylases suggest roles for H3K4me1 in transcription. However, the control of H3K4me1 remains unexplored in Arabidopsis, in which no methyltransferase for H3K4me1 has been identified. Here, we identify three Arabidopsis methyltransferases that direct H3K4me1. Analyses of their genome-wide localization using ChIP-seq and machine learning reveal that one of the enzymes cooperates with the transcription machinery, while the other two are associated with specific histone modifications and DNA sequences. Importantly, these two types of localization patterns are also found for the other H3K4 methyltransferases in Arabidopsis and mice. These results suggest that H3K4me1/2/3 are established and maintained via interplay with transcription as well as inputs from other chromatin features, presumably enabling elaborate gene control.
    DOI:  https://doi.org/10.1038/s41467-022-32165-8
  15. Cells. 2022 Aug 03. pii: 2392. [Epub ahead of print]11(15):
    Endogenous Retroviral Sequences Behave as Putative Enhancers Controlling Gene Expression through HP1-Regulated Long-Range Chromatin Interactions.
    Sébastien Calvet, Séphora Sallis, Nehmé Saksouk, Cosette Rebouissou, Catherine Teyssier, Annick Lesne, Florence Cammas, Thierry Forné.
      About half of the mammalian genome is constituted of repeated elements, among which endogenous retroviruses (ERVs) are known to influence gene expression and cancer development. The HP1 (Heterochromatin Protein 1) proteins are known to be essential for heterochromatin establishment and function and its loss in hepatocytes leads to the reactivation of specific ERVs and to liver tumorigenesis. Here, by studying two ERVs located upstream of genes upregulated upon loss of HP1, Mbd1 and Trim24, we show that these HP1-dependent ERVs behave as either alternative promoters or as putative enhancers forming a loop with promoters of endogenous genes depending on the genomic context and HP1 expression level. These ERVs are characterised by a specific HP1-independent enrichment in heterochromatin-associated marks H3K9me3 and H4K20me3 as well as in the enhancer-specific mark H3K4me1, a combination that might represent a bookmark of putative ERV-derived enhancers. These ERVs are further enriched in a HP1-dependent manner in H3K27me3, suggesting a critical role of this mark together with HP1 in the silencing of the ERVs, as well as for the repression of the associated genes. Altogether, these results lead to the identification of a new regulatory hub involving the HP1-dependent formation of a physical loop between specific ERVs and endogenous genes.
    Keywords:  HP1; Trim24; chromatin organization; endogenous retroviruses
    DOI:  https://doi.org/10.3390/cells11152392
  16. Bio Protoc. 2022 Jul 05. pii: e4460. [Epub ahead of print]12(13):
    HaloChIP-seq for Antibody-Independent Mapping of Mouse Transcription Factor Cistromes in vivo.
    Ann Louise Hunter, Antony D Adamson, Toryn M Poolman, Magdalena Grudzien, Andrew S I Loudon, David W Ray, David A Bechtold.
      Chromatin immunoprecipitation (ChIP) maps, on a genome-wide scale, transcription factor binding sites, and the distribution of other chromatin-associated proteins and their modifications. As such, it provides valuable insights into mechanisms of gene regulation. However, successful ChIP experiments are dependent on the availability of a high-quality antibody against the target of interest. Using antibodies with poor sensitivity and specificity can yield misleading results. This can be partly circumvented by using epitope-tagged systems ( e.g. , HA, Myc, His), but these approaches are still antibody-dependent. HaloTag ® is a modified dehalogenase enzyme, which covalently binds synthetic ligands. This system can be used for imaging and purification of HaloTag ® fusion proteins, and has been used for ChIP in vitro . Here, we present a protocol for using the HaloTag ® system for ChIP in vivo , to map, with sensitivity and specificity, the cistrome of a dynamic mouse transcription factor expressed at its endogenous locus. Graphical abstract.
    Keywords:   ChIP ; Fusion protein ; HaloTag ® ; NR1D1 ; Nuclear receptor ; Tag
    DOI:  https://doi.org/10.21769/BioProtoc.4460
  17. Proc Natl Acad Sci U S A. 2022 Aug 16. 119(33): e2204338119
    Adrenergic receptor signaling induced by Klf15, a regulator of regeneration enhancer, promotes kidney reconstruction.
    Nanoka Suzuki, Akinori Kanai, Yutaka Suzuki, Hajime Ogino, Haruki Ochi.
      Despite the recent discovery of tissue regeneration enhancers in highly regenerative animals, upstream and downstream genetic programs connected by these enhancers still remain unclear. Here, we performed a genome-wide analysis of enhancers and associated genes in regenerating nephric tubules of Xenopus laevis. Putative enhancers were identified using assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) and H3K27ac chromatin immunoprecipitation sequencing (ChIP-seq) analyses. Their target genes were predicted based on their proximity to enhancers on genomic DNA and consistency of their transcriptome profiles to ATAC-seq/ChIP-seq profiles of the enhancers. Motif enrichment analysis identified the central role of Krüppel-like factors (Klf) in the enhancer. Klf15, a member of the Klf family, directly binds enhancers and stimulates expression of regenerative genes, including adrenoreceptor alpha 1A (adra1a), whereas inhibition of Klf15 activity results in failure of nephric tubule regeneration. Moreover, pharmacological inhibition of Adra1a-signaling suppresses nephric tubule regeneration, while its activation promotes nephric tubule regeneration and restores organ size. These results indicate that Klf15-dependent adrenergic receptor signaling through regeneration enhancers plays a central role in the genetic network for kidney regeneration.
    Keywords:  Klf transcription factor; Xenopus laevis; adrenergic receptor; kidney; regeneration enhancer
    DOI:  https://doi.org/10.1073/pnas.2204338119
  18. PLoS Comput Biol. 2022 Aug;18(8): e1010393
    Chromatin accessibility of primary human cancers ties regional mutational processes and signatures with tissues of origin.
    Oliver Ocsenas, Jüri Reimand.
      Somatic mutations in cancer genomes are associated with DNA replication timing (RT) and chromatin accessibility (CA), however these observations are based on normal tissues and cell lines while primary cancer epigenomes remain uncharacterised. Here we use machine learning to model megabase-scale mutation burden in 2,500 whole cancer genomes and 17 cancer types via a compendium of 900 CA and RT profiles covering primary cancers, normal tissues, and cell lines. CA profiles of primary cancers, rather than those of normal tissues, are most predictive of regional mutagenesis in most cancer types. Feature prioritisation shows that the epigenomes of matching cancer types and organ systems are often the strongest predictors of regional mutation burden, highlighting disease-specific associations of mutational processes. The genomic distributions of mutational signatures are also shaped by the epigenomes of matched cancer and tissue types, with SBS5/40, carcinogenic and unknown signatures most accurately predicted by our models. In contrast, fewer associations of RT and regional mutagenesis are found. Lastly, the models highlight genomic regions with overrepresented mutations that dramatically exceed epigenome-derived expectations and show a pan-cancer convergence to genes and pathways involved in development and oncogenesis, indicating the potential of this approach for coding and non-coding driver discovery. The association of regional mutational processes with the epigenomes of primary cancers suggests that the landscape of passenger mutations is predominantly shaped by the epigenomes of cancer cells after oncogenic transformation.
    DOI:  https://doi.org/10.1371/journal.pcbi.1010393
  19. Brief Bioinform. 2022 Aug 12. pii: bbac329. [Epub ahead of print]
    GREAP: a comprehensive enrichment analysis software for human genomic regions.
    Yongsan Yang, Fengcui Qian, Xuecang Li, Yanyu Li, Liwei Zhou, Qiuyu Wang, Xinyuan Zhou, Jian Zhang, Chao Song, Zhengmin Yu, Ting Cui, Chenchen Feng, Jiang Zhu, Desi Shang, Jiaqi Liu, Mengfei Sun, Yuexin Zhang, Huifang Tang, Chunquan Li.
      The rapid development of genomic high-throughput sequencing has identified a large number of DNA regulatory elements with abundant epigenetics markers, which promotes the rapid accumulation of functional genomic region data. The comprehensively understanding and research of human functional genomic regions is still a relatively urgent work at present. However, the existing analysis tools lack extensive annotation and enrichment analytical abilities for these regions. Here, we designed a novel software, Genomic Region sets Enrichment Analysis Platform (GREAP), which provides comprehensive region annotation and enrichment analysis capabilities. Currently, GREAP supports 85 370 genomic region reference sets, which cover 634 681 107 regions across 11 different data types, including super enhancers, transcription factors, accessible chromatins, etc. GREAP provides widespread annotation and enrichment analysis of genomic regions. To reflect the significance of enrichment analysis, we used the hypergeometric test and also provided a Locus Overlap Analysis. In summary, GREAP is a powerful platform that provides many types of genomic region sets for users and supports genomic region annotations and enrichment analyses. In addition, we developed a customizable genome browser containing >400 000 000 customizable tracks for visualization. The platform is freely available at http://www.liclab.net/Greap/view/index.
    Keywords:  deep annotation; enrichment analysis; epigenetics; genomic region
    DOI:  https://doi.org/10.1093/bib/bbac329
  20. Elife. 2022 Aug 12. pii: e71591. [Epub ahead of print]11
    Generation of functional hepatocytes by forward programming with nuclear receptors.
    Rute A Tomaz, Ekaterini D Zacharis, Fabian Bachinger, Annabelle Wurmser, Daniel Yamamoto, Sandra Petrus-Reurer, Carola M Morell, Dominika Dziedzicka, Brandon T Wesley, Imbisaat Geti, Charis-Patricia Segeritz, Miguel C de Brito, Mariya Chhatriwala, Daniel Ortmann, Kourosh Saeb-Parsy, Ludovic Vallier.
      Production of large quantities of hepatocytes remains a major challenge for a number of clinical applications in the biomedical field. Directed differentiation of human pluripotent stem cells (hPSCs) into hepatocyte-like cells (HLCs) provides an advantageous solution and a number of protocols have been developed for this purpose. However, these methods usually follow different steps of liver development in vitro, which is time consuming and requires complex culture conditions. In addition, HLCs lack the full repertoire of functionalities characterising primary hepatocytes. Here, we explore the interest of forward programming to generate hepatocytes from hPSCs and to bypass these limitations. This approach relies on the overexpression of three hepatocyte nuclear factors (HNF1A, HNF6, and FOXA3) in combination with different nuclear receptors expressed in the adult liver using the OPTi-OX platform. Forward programming allows for the rapid production of hepatocytes (FoP-Heps) with functional characteristics using a simplified process. We also uncovered that the overexpression of nuclear receptors such as RORc can enhance specific functionalities of FoP-Heps thereby validating its role in lipid/glucose metabolism. Together, our results show that forward programming could offer a versatile alternative to direct differentiation for generating hepatocytes in vitro.
    Keywords:  forward programming; hepatocytes; human; pluripotent stem cells; regenerative medicine; stem cells
    DOI:  https://doi.org/10.7554/eLife.71591
  21. Genetics. 2022 Aug 08. pii: iyac111. [Epub ahead of print]
    Transcription factors perform a 2-step search of the nucleus.
    Max Valentín Staller.
      Transcription factors regulate gene expression by binding to regulatory DNA and recruiting regulatory protein complexes. The DNA-binding and protein-binding functions of transcription factors are traditionally described as independent functions performed by modular protein domains. Here, I argue that genome binding can be a 2-part process with both DNA-binding and protein-binding steps, enabling transcription factors to perform a 2-step search of the nucleus to find their appropriate binding sites in a eukaryotic genome. I support this hypothesis with new and old results in the literature, discuss how this hypothesis parsimoniously resolves outstanding problems, and present testable predictions.
    Keywords:  DNA-binding domain; gene regulation; intrinsically disordered region; liquid–liquid phase separation; transcription; transcription factor; transcription initiation; transcriptional activation domain; transcriptional condensate
    DOI:  https://doi.org/10.1093/genetics/iyac111
  22. Nucleic Acids Res. 2022 Aug 10. pii: gkac678. [Epub ahead of print]
    Differential dependencies of human RNA polymerase II promoters on TBP, TAF1, TFIIB and XPB.
    Juan F Santana, Geoffrey S Collins, Mrutyunjaya Parida, Donal S Luse, David H Price.
      The effects of rapid acute depletion of components of RNA polymerase II (Pol II) general transcription factors (GTFs) that are thought to be critical for formation of preinitiation complexes (PICs) and initiation in vitro were quantified in HAP1 cells using precision nuclear run-on sequencing (PRO-Seq). The average dependencies for each factor across >70 000 promoters varied widely even though levels of depletions were similar. Some of the effects could be attributed to the presence or absence of core promoter elements such as the upstream TBP-specificity motif or downstream G-rich sequences, but some dependencies anti-correlated with such sequences. While depletion of TBP had a large effect on most Pol III promoters only a small fraction of Pol II promoters were similarly affected. TFIIB depletion had the largest general effect on Pol II and also correlated with apparent termination defects downstream of genes. Our results demonstrate that promoter activity is combinatorially influenced by recruitment of TFIID and sequence-specific transcription factors. They also suggest that interaction of the preinitiation complex (PIC) with nucleosomes can affect activity and that recruitment of TFIID containing TBP only plays a positive role at a subset of promoters.
    DOI:  https://doi.org/10.1093/nar/gkac678
  23. Cell Death Discov. 2022 Aug 12. 8(1): 356
    FTO promotes clear cell renal cell carcinoma progression via upregulation of PDK1 through an m6A dependent pathway.
    Haixiang Shen, Yufan Ying, Xueyou Ma, Haiyun Xie, Shiming Chen, Jiazhu Sun, Zixiang Liu, Chao Wen, Zitong Yang, Xiao Wang, Mingjie Xu, Jindan Luo, Ben Liu, Jiangfeng Li, Xiangyi Zheng, Liping Xie.
      FTO, as an m6A mRNA demethylase, is involved in various cancers. However, the role of FTO in clear cell renal cell carcinoma (ccRCC) remains unclear. In the present study, we discovered FTO is upregulated in ccRCC. Functionally, knockdown of FTO significantly impairs the proliferation and migration ability of ccRCC cells. Mechanistically, our data suggest FTO promotes the proliferation and migration of ccRCC through preventing degradation of PDK1 mRNA induced by YTHDF2 in an m6A-dependent mechanism. Overall, our results identify the protumorigenic role of FTO through the m6A/YTHDF2/PDK1 pathway, which could be a promising therapeutic target for ccRCC.
    DOI:  https://doi.org/10.1038/s41420-022-01151-w
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