bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2021‒07‒04
twenty-one papers selected by
Connor Rogerson
University of Cambridge, MRC Cancer Unit
  1. Histone H3K4me1 strongly activates the DNase I hypersensitive sites in super-enhancers than those in typical enhancers.
  2. Interplay between CTCF boundaries and a super enhancer controls cohesin extrusion trajectories and gene expression.
  3. Orphan CpG islands amplify poised enhancer regulatory activity and determine target gene responsiveness.
  4. Pax7 pioneer factor action requires both paired and homeo DNA binding domains.
  5. BAP1 enhances Polycomb repression by counteracting widespread H2AK119ub1 deposition and chromatin condensation.
  6. Systematic dissection of transcriptional regulatory networks by genome-scale and single-cell CRISPR screens.
  7. UniBind: maps of high-confidence direct TF-DNA interactions across nine species.
  8. AP-1 activity is a major barrier of human somatic cell reprogramming.
  9. Phosphorylation of the HP1β hinge region sequesters KAP1 in heterochromatin and promotes the exit from naïve pluripotency.
  10. scRNA-Sequencing uncovers a TCF4-dependent transcription factor network regulating commissure development.
  11. AP-2alpha-mediated activation of E2F and EZH2 drives melanoma metastasis.
  12. Extended and dynamic linker histone-DNA Interactions control chromatosome compaction.
  13. Oxygen concentration affects de novo DNA methylation and transcription in in vitro cultured oocytes.
  14. NEUROD1 Is Required for the Early α and β Endocrine Differentiation in the Pancreas.
  15. Genome-Wide Histone H3K27 Acetylation Profiling Identified Genes Correlated With Prognosis in Papillary Thyroid Carcinoma.
  16. A functional map of genomic HIF1α-DNA complexes in the eye lens revealed through multiomics analysis.
  17. Cell-free genomics reveal intrinsic, cooperative and competitive determinants of chromatin interactions.
  18. Genetic drivers of m6A methylation in human brain, lung, heart and muscle.
  19. A Bayesian inference transcription factor activity model for the analysis of single-cell transcriptomes.
  20. Forkhead box family transcription factors as versatile regulators for cellular reprogramming to pluripotency.
  21. Direct phosphorylation and stabilization of HIF-1α by PIM1 kinase drives angiogenesis in solid tumors.
  1. Biosci Rep. 2021 Jun 25. pii: BSR20210691. [Epub ahead of print]
    Histone H3K4me1 strongly activates the DNase I hypersensitive sites in super-enhancers than those in typical enhancers.
    Yujin Kang, Jin Kang, AeRi Kim.
      Super-enhancers, which consist of multiple enhancer elements, are occupied by master transcription factors and coactivators, such as Mediator, and are highly acetylated at histone H3K27. Here, we have characterized the super-enhancers in terms of DNase I hypersensitive sites (DHSs) by analyzing publicly available ChIP-seq and DNase-seq data of K562 cells and compared to the DHSs in typical enhancers. DHSs in the super-enhancers were highly marked by histone H3K4me1 than DHSs in typical enhancers. Loss of H3K4me1 by the deletion of catalytic domains in histone methyltransferases MLL3 and MLL4 remarkably decreased histone H3K27ac and histone H3 depletion at super-enhancer DHSs than at typical enhancer DHSs. The levels of enhancer RNA (eRNA) transcripts and mRNA transcripts from the putative target genes were notably reduced at and near super-enhancer DHSs than typical enhancer DHSs following H3K4me1 loss. These results indicate that histone H3K4me1 is a marker for DHSs in super-enhancers and that this modification has a more significant impact on the activation of super-enhancer DHSs than typical enhancer DHSs.
    Keywords:  DNase I hypersensitive site; H3K27ac; H3K4me1; Super-enhancer; eRNA; histone depletion
    DOI:  https://doi.org/10.1042/BSR20210691
  2. Mol Cell. 2021 Jun 22. pii: S1097-2765(21)00455-X. [Epub ahead of print]
    Interplay between CTCF boundaries and a super enhancer controls cohesin extrusion trajectories and gene expression.
    Erica S M Vos, Christian Valdes-Quezada, Yike Huang, Amin Allahyar, Marjon J A M Verstegen, Anna-Karina Felder, Floor van der Vegt, Esther C H Uijttewaal, Peter H L Krijger, Wouter de Laat.
      To understand how chromatin domains coordinate gene expression, we dissected select genetic elements organizing topology and transcription around the Prdm14 super enhancer in mouse embryonic stem cells. Taking advantage of allelic polymorphisms, we developed methods to sensitively analyze changes in chromatin topology, gene expression, and protein recruitment. We show that enhancer insulation does not rely strictly on loop formation between its flanking boundaries, that the enhancer activates the Slco5a1 gene beyond its prominent domain boundary, and that it recruits cohesin for loop extrusion. Upon boundary inversion, we find that oppositely oriented CTCF terminates extrusion trajectories but does not stall cohesin, while deleted or mutated CTCF sites allow cohesin to extend its trajectory. Enhancer-mediated gene activation occurs independent of paused loop extrusion near the gene promoter. We expand upon the loop extrusion model to propose that cohesin loading and extrusion trajectories originating at an enhancer contribute to gene activation.
    Keywords:  CTCF; TADs; chromatin; cohesin; domains; enhancers; gene regulation; genome organization; loop extrusion; loops
    DOI:  https://doi.org/10.1016/j.molcel.2021.06.008
  3. Nat Genet. 2021 Jun 28.
    Orphan CpG islands amplify poised enhancer regulatory activity and determine target gene responsiveness.
    Tomas Pachano, Víctor Sánchez-Gaya, Thais Ealo, Maria Mariner-Faulí, Tore Bleckwehl, Helena G Asenjo, Patricia Respuela, Sara Cruz-Molina, María Muñoz-San Martín, Endika Haro, Wilfred F J van IJcken, David Landeira, Alvaro Rada-Iglesias.
      CpG islands (CGIs) represent a widespread feature of vertebrate genomes, being associated with ~70% of all gene promoters. CGIs control transcription initiation by conferring nearby promoters with unique chromatin properties. In addition, there are thousands of distal or orphan CGIs (oCGIs) whose functional relevance is barely known. Here we show that oCGIs are an essential component of poised enhancers that augment their long-range regulatory activity and control the responsiveness of their target genes. Using a knock-in strategy in mouse embryonic stem cells, we introduced poised enhancers with or without oCGIs within topologically associating domains harboring genes with different types of promoters. Analysis of the resulting cell lines revealed that oCGIs act as tethering elements that promote the physical and functional communication between poised enhancers and distally located genes, particularly those with large CGI clusters in their promoters. Therefore, by acting as genetic determinants of gene-enhancer compatibility, CGIs can contribute to gene expression control under both physiological and potentially pathological conditions.
    DOI:  https://doi.org/10.1038/s41588-021-00888-x
  4. Nucleic Acids Res. 2021 Jul 01. pii: gkab561. [Epub ahead of print]
    Pax7 pioneer factor action requires both paired and homeo DNA binding domains.
    Audrey Pelletier, Alexandre Mayran, Arthur Gouhier, James G Omichinski, Aurelio Balsalobre, Jacques Drouin.
      The pioneer transcription factor Pax7 contains two DNA binding domains (DBD), a paired and a homeo domain. Previous work on Pax7 and the related Pax3 showed that each DBD binds a cognate DNA sequence, thus defining two targets of binding and possibly modalities of action. Genomic targets of Pax7 pioneer action leading to chromatin opening are enriched for composite DNA target sites containing juxtaposed sites for both paired and homeo domains. The present work investigated the implication of the DBDs in pioneer action. We show that the composite sequence is a higher affinity binding site and that efficient binding to this site involves both DBDs of the same Pax7 molecule. This binding is not sensitive to cytosine methylation of the DNA sites consistent with pioneer action within nucleosomal heterochromatin. Introduction of single amino acid mutations in either paired or homeo domain that impair binding to cognate DNA sequences showed that both DBDs must be intact for pioneer action. In contrast, only the paired domain is required for low affinity binding of heterochromatin sites. Thus, Pax7 pioneer action on heterochromatin requires unique protein:DNA interactions that are more complex compared to its simpler DNA binding modalities at accessible enhancer target sites.
    DOI:  https://doi.org/10.1093/nar/gkab561
  5. Mol Cell. 2021 Jun 24. pii: S1097-2765(21)00500-1. [Epub ahead of print]
    BAP1 enhances Polycomb repression by counteracting widespread H2AK119ub1 deposition and chromatin condensation.
    Eric Conway, Federico Rossi, Daniel Fernandez-Perez, Eleonora Ponzo, Karin Johanna Ferrari, Marika Zanotti, Daria Manganaro, Simona Rodighiero, Simone Tamburri, Diego Pasini.
      BAP1 is mutated or deleted in many cancer types, including mesothelioma, uveal melanoma, and cholangiocarcinoma. It is the catalytic subunit of the PR-DUB complex, which removes PRC1-mediated H2AK119ub1, essential for maintaining transcriptional repression. However, the precise relationship between BAP1 and Polycombs remains elusive. Using embryonic stem cells, we show that BAP1 restricts H2AK119ub1 deposition to Polycomb target sites. This increases the stability of Polycomb with their targets and prevents diffuse accumulation of H2AK119ub1 and H3K27me3. Loss of BAP1 results in a broad increase in H2AK119ub1 levels that is primarily dependent on PCGF3/5-PRC1 complexes. This titrates PRC2 away from its targets and stimulates H3K27me3 accumulation across the genome, leading to a general chromatin compaction. This provides evidence for a unifying model that resolves the apparent contradiction between BAP1 catalytic activity and its role in vivo, uncovering molecular vulnerabilities that could be useful for BAP1-related pathologies.
    Keywords:  BAP1; H2AK119ub1; PCGF3; PR-DUB; PRC1; PRC2; Polycomb; chromatin compaction; transcriptional repression; tumor suppressor
    DOI:  https://doi.org/10.1016/j.molcel.2021.06.020
  6. Sci Adv. 2021 Jul;pii: eabf5733. [Epub ahead of print]7(27):
    Systematic dissection of transcriptional regulatory networks by genome-scale and single-cell CRISPR screens.
    Rui Lopes, Kathleen Sprouffske, Caibin Sheng, Esther C H Uijttewaal, Adriana Emma Wesdorp, Jan Dahinden, Simon Wengert, Juan Diaz-Miyar, Umut Yildiz, Melusine Bleu, Verena Apfel, Fanny Mermet-Meillon, Rok Krese, Mathias Eder, André Vidas Olsen, Philipp Hoppe, Judith Knehr, Walter Carbone, Rachel Cuttat, Annick Waldt, Marc Altorfer, Ulrike Naumann, Joachim Weischenfeldt, Antoine deWeck, Audrey Kauffmann, Guglielmo Roma, Dirk Schübeler, Giorgio G Galli.
      Millions of putative transcriptional regulatory elements (TREs) have been cataloged in the human genome, yet their functional relevance in specific pathophysiological settings remains to be determined. This is critical to understand how oncogenic transcription factors (TFs) engage specific TREs to impose transcriptional programs underlying malignant phenotypes. Here, we combine cutting edge CRISPR screens and epigenomic profiling to functionally survey ≈15,000 TREs engaged by estrogen receptor (ER). We show that ER exerts its oncogenic role in breast cancer by engaging TREs enriched in GATA3, TFAP2C, and H3K27Ac signal. These TREs control critical downstream TFs, among which TFAP2C plays an essential role in ER-driven cell proliferation. Together, our work reveals novel insights into a critical oncogenic transcription program and provides a framework to map regulatory networks, enabling to dissect the function of the noncoding genome of cancer cells.
    DOI:  https://doi.org/10.1126/sciadv.abf5733
  7. BMC Genomics. 2021 Jun 26. 22(1): 482
    UniBind: maps of high-confidence direct TF-DNA interactions across nine species.
    Rafael Riudavets Puig, Paul Boddie, Aziz Khan, Jaime Abraham Castro-Mondragon, Anthony Mathelier.
      BACKGROUND: Transcription factors (TFs) bind specifically to TF binding sites (TFBSs) at cis-regulatory regions to control transcription. It is critical to locate these TF-DNA interactions to understand transcriptional regulation. Efforts to predict bona fide TFBSs benefit from the availability of experimental data mapping DNA binding regions of TFs (chromatin immunoprecipitation followed by sequencing - ChIP-seq).RESULTS: In this study, we processed ~ 10,000 public ChIP-seq datasets from nine species to provide high-quality TFBS predictions. After quality control, it culminated with the prediction of ~ 56 million TFBSs with experimental and computational support for direct TF-DNA interactions for 644 TFs in > 1000 cell lines and tissues. These TFBSs were used to predict > 197,000 cis-regulatory modules representing clusters of binding events in the corresponding genomes. The high-quality of the TFBSs was reinforced by their evolutionary conservation, enrichment at active cis-regulatory regions, and capacity to predict combinatorial binding of TFs. Further, we confirmed that the cell type and tissue specificity of enhancer activity was correlated with the number of TFs with binding sites predicted in these regions. All the data is provided to the community through the UniBind database that can be accessed through its web-interface ( https://unibind.uio.no/ ), a dedicated RESTful API, and as genomic tracks. Finally, we provide an enrichment tool, available as a web-service and an R package, for users to find TFs with enriched TFBSs in a set of provided genomic regions.
    CONCLUSIONS: UniBind is the first resource of its kind, providing the largest collection of high-confidence direct TF-DNA interactions in nine species.
    Keywords:  ChIP-seq; Cis-regulatory modules; Evolutionary conservation; TF-DNA interactions; Transcription factor binding sites; Transcription regulation; UniBind
    DOI:  https://doi.org/10.1186/s12864-021-07760-6
  8. Cell Mol Life Sci. 2021 Jun 28.
    AP-1 activity is a major barrier of human somatic cell reprogramming.
    Yuting Liu, Jiangping He, Ruhai Chen, He Liu, Jocelyn Chen, Yujian Liu, Bo Wang, Lin Guo, Duanqing Pei, Jie Wang, Jing Liu, Jiekai Chen.
      Human induced pluripotent stem cells (iPSCs) technology has been widely applied to cell regeneration and disease modeling. However, most mechanism of somatic reprogramming is studied on mouse system, which is not always generic in human. Consequently, the generation of human iPSCs remains inefficient. Here, we map the chromatin accessibility dynamics during the induction of human iPSCs from urine cells. Comparing to the mouse system, we found that the closing of somatic loci is much slower in human. Moreover, a conserved AP-1 motif is highly enriched among the closed loci. The introduction of AP-1 repressor, JDP2, enhances human reprogramming and facilitates the reactivation of pluripotent genes. However, ESRRB, KDM2B and SALL4, several known pluripotent factors promoting mouse somatic reprogramming fail to enhance human iPSC generation. Mechanistically, we reveal that JDP2 promotes the closing of somatic loci enriching AP-1 motifs to enhance human reprogramming. Furthermore, JDP2 can rescue reprogramming deficiency without MYC or KLF4. These results indicate AP-1 activity is a major barrier to prevent chromatin remodeling during somatic cell reprogramming.
    Keywords:  AP-1; Chromatin remodeling; Human induced pluripotent stem cells; JDP2
    DOI:  https://doi.org/10.1007/s00018-021-03883-x
  9. Nucleic Acids Res. 2021 Jul 02. pii: gkab548. [Epub ahead of print]
    Phosphorylation of the HP1β hinge region sequesters KAP1 in heterochromatin and promotes the exit from naïve pluripotency.
    Weihua Qin, Enes Ugur, Christopher B Mulholland, Sebastian Bultmann, Irina Solovei, Miha Modic, Martha Smets, Michael Wierer, Ignasi Forné, Axel Imhof, M Cristina Cardoso, Heinrich Leonhardt.
      Heterochromatin binding protein HP1β plays an important role in chromatin organization and cell differentiation, however the underlying mechanisms remain unclear. Here, we generated HP1β-/- embryonic stem cells and observed reduced heterochromatin clustering and impaired differentiation. We found that during stem cell differentiation, HP1β is phosphorylated at serine 89 by CK2, which creates a binding site for the pluripotency regulator KAP1. This phosphorylation dependent sequestration of KAP1 in heterochromatin compartments causes a downregulation of pluripotency factors and triggers pluripotency exit. Accordingly, HP1β-/- and phospho-mutant cells exhibited impaired differentiation, while ubiquitination-deficient KAP1-/- cells had the opposite phenotype with enhanced differentiation. These results suggest that KAP1 regulates pluripotency via its ubiquitination activity. We propose that the formation of subnuclear membraneless heterochromatin compartments may serve as a dynamic reservoir to trap or release cellular factors. The sequestration of essential regulators defines a novel and active role of heterochromatin in gene regulation and represents a dynamic mode of remote control to regulate cellular processes like cell fate decisions.
    DOI:  https://doi.org/10.1093/nar/gkab548
  10. Development. 2021 Jun 29. pii: dev.196022. [Epub ahead of print]
    scRNA-Sequencing uncovers a TCF4-dependent transcription factor network regulating commissure development.
    Marie-Theres Wittmann, Sayako Katada, Elisabeth Sock, Philipp Kirchner, Arif B Ekici, Michael Wegner, Kinichi Nakashima, D Chichung Lie, André Reis.
      Transcription factor 4 (TCF4) is a critical regulator of neurodevelopment and has been linked to the pathogenesis of autism, intellectual disability, and schizophrenia. As a class I bHLH transcription factor it is assumed that TCF4 exerts its neurodevelopmental functions through dimerization with proneural class II bHLH TFs. Here, we aim to identify transcription factor (TF) partners of TCF4 in the control of interhemispheric connectivity formation. Using a new bioinformatic strategy integrating TF expression levels and regulon activities from single cell RNA-sequencing data, we find evidence that TCF4 interacts with non-bHLH TFs and modulates their transcriptional activity in Satb2+ intercortical projection neurons. Notably, this network comprises regulators linked to the pathogenesis of neurodevelopmental disorders, e.g. FOXG1, SOX11 and BRG1. In support of the functional interaction of TCF4 with non-bHLH TFs we find that TCF4 and SOX11 biochemically interact and cooperatively control commissure formation in vivo, and regulate the transcription of genes implied in this process. Next to identifying new candidate interactors of TCF4 in neurodevelopment, this study illustrates how scRNA-Seq data can be leveraged to predict TFs networks in neurodevelopmental processes.
    Keywords:  Commissure development; Gene regulatory networks; Protein-protein interaction; SOX11; Single-cell RNA-Sequencing; TCF4
    DOI:  https://doi.org/10.1242/dev.196022
  11. Cancer Res. 2021 Jul 01. pii: canres.0772.2021. [Epub ahead of print]
    AP-2alpha-mediated activation of E2F and EZH2 drives melanoma metastasis.
    Jeffrey R White, Dakota T Thompson, Kelsey E Koch, Boris S Kiriazov, Anna C Beck, Dana M van der Heide, Benjamin G Grimm, Mikhail V Kulak, Ronald J Weigel.
      In melanoma metastasis, the role of the AP-2alpha transcription factor, which is encoded by TFAP2A, is controversial as some findings have suggested tumor suppressor activity while other studies have shown high TFAP2A expression in node-positive melanoma associated with poor prognosis. Here we demonstrate that AP-2alpha facilitates melanoma metastasis through transcriptional activation of genes within the E2F pathway including EZH2. A BioID screen found that AP-2alpha interacts with members of the nucleosome remodeling and deacetylase (NuRD) complex. Loss of AP-2alpha removed activating chromatin marks in the promoters of EZH2 and other E2F target genes through activation of the NuRD repression complex. In melanoma cells, treatment with tazemetostat, an FDA-approved and highly specific EZH2 inhibitor, substantially reduced anchorage-independent colony formation and demonstrated heritable anti-metastatic effects, which were dependent on AP-2alpha. Single cell RNA-seq analysis of a metastatic melanoma mouse model revealed hyperexpansion of Tfap2aHigh/E2F-activated cell populations in transformed melanoma relative to progenitor melanocyte stem cells. These findings demonstrate that melanoma metastasis is driven by the AP-2alpha/EZH2 pathway and suggest that AP-2alpha expression can be used as a biomarker to predict responsiveness to EZH2 inhibitors for the treatment of advanced melanomas.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-21-0772
  12. Mol Cell. 2021 Jun 19. pii: S1097-2765(21)00453-6. [Epub ahead of print]
    Extended and dynamic linker histone-DNA Interactions control chromatosome compaction.
    Sergei Rudnizky, Hadeel Khamis, Yuval Ginosar, Efrat Goren, Philippa Melamed, Ariel Kaplan.
      Chromatosomes play a fundamental role in chromatin regulation, but a detailed understanding of their structure is lacking, partially due to their complex dynamics. Using single-molecule DNA unzipping with optical tweezers, we reveal that linker histone interactions with DNA are remarkably extended, with the C-terminal domain binding both DNA linkers as far as approximately ±140 bp from the dyad. In addition to a symmetrical compaction of the nucleosome core governed by globular domain contacts at the dyad, the C-terminal domain compacts the nucleosome's entry and exit. These interactions are dynamic, exhibit rapid binding and dissociation, are sensitive to phosphorylation of a specific residue, and are crucial to determining the symmetry of the chromatosome's core. Extensive unzipping of the linker DNA, which mimics its invasion by motor proteins, shifts H1 into an asymmetric, off-dyad configuration and triggers nucleosome decompaction, highlighting the plasticity of the chromatosome structure and its potential regulatory role.
    Keywords:  DNA unzipping; chromatin; chromatosome; compaction; linker histone; nucleosome; optical tweezers; single-molecule biophysics
    DOI:  https://doi.org/10.1016/j.molcel.2021.06.006
  13. Clin Epigenetics. 2021 Jun 28. 13(1): 132
    Oxygen concentration affects de novo DNA methylation and transcription in in vitro cultured oocytes.
    Florence Naillat, Heba Saadeh, Joanna Nowacka-Woszuk, Lenka Gahurova, Fatima Santos, Shin-Ichi Tomizawa, Gavin Kelsey.
      BACKGROUND: Reproductive biology methods rely on in vitro follicle cultures from mature follicles obtained by hormonal stimulation for generating metaphase II oocytes to be fertilised and developed into a healthy embryo. Such techniques are used routinely in both rodent and human species. DNA methylation is a dynamic process that plays a role in epigenetic regulation of gametogenesis and development. In mammalian oocytes, DNA methylation establishment regulates gene expression in the embryos. This regulation is particularly important for a class of genes, imprinted genes, whose expression patterns are crucial for the next generation. The aim of this work was to establish an in vitro culture system for immature mouse oocytes that will allow manipulation of specific factors for a deeper analysis of regulatory mechanisms for establishing transcription regulation-associated methylation patterns.RESULTS: An in vitro culture system was developed from immature mouse oocytes that were grown to germinal vesicles (GV) under two different conditions: normoxia (20% oxygen, 20% O2) and hypoxia (5% oxygen, 5% O2). The cultured oocytes were sorted based on their sizes. Reduced representative bisulphite sequencing (RRBS) and RNA-seq libraries were generated from cultured and compared to in vivo-grown oocytes. In the in vitro cultured oocytes, global and CpG-island (CGI) methylation increased gradually along with oocyte growth, and methylation of the imprinted genes was similar to in vivo-grown oocytes. Transcriptomes of the oocytes grown in normoxia revealed chromatin reorganisation and enriched expression of female reproductive genes, whereas in the 5% O2 condition, transcripts were biased towards cellular stress responses. To further confirm the results, we developed a functional assay based on our model for characterising oocyte methylation using drugs that reduce methylation and transcription. When histone methylation and transcription processes were reduced, DNA methylation at CGIs from gene bodies of grown oocytes presented a lower methylation profile.
    CONCLUSIONS: Our observations reveal changes in DNA methylation and transcripts between oocytes cultured in vitro with different oxygen concentrations and in vivo-grown murine oocytes. Oocytes grown under 20% O2 had a higher correlation with in vivo oocytes for DNA methylation and transcription demonstrating that higher oxygen concentration is beneficial for the oocyte maturation in ex vivo culture condition. Our results shed light on epigenetic mechanisms for the development of oocytes from an immature to GV oocyte in an in vitro culture model.
    Keywords:  5% oxygen; DNA methylation; In vitro culture; Mouse; Normoxia; Oocyte; Transcription
    DOI:  https://doi.org/10.1186/s13148-021-01116-3
  14. Int J Mol Sci. 2021 Jun 23. pii: 6713. [Epub ahead of print]22(13):
    NEUROD1 Is Required for the Early α and β Endocrine Differentiation in the Pancreas.
    Romana Bohuslavova, Ondrej Smolik, Jessica Malfatti, Zuzana Berkova, Zaneta Novakova, Frantisek Saudek, Gabriela Pavlinkova.
      Diabetes is a metabolic disease that involves the death or dysfunction of the insulin-secreting β cells in the pancreas. Consequently, most diabetes research is aimed at understanding the molecular and cellular bases of pancreatic development, islet formation, β-cell survival, and insulin secretion. Complex interactions of signaling pathways and transcription factor networks regulate the specification, growth, and differentiation of cell types in the developing pancreas. Many of the same regulators continue to modulate gene expression and cell fate of the adult pancreas. The transcription factor NEUROD1 is essential for the maturation of β cells and the expansion of the pancreatic islet cell mass. Mutations of the Neurod1 gene cause diabetes in humans and mice. However, the different aspects of the requirement of NEUROD1 for pancreas development are not fully understood. In this study, we investigated the role of NEUROD1 during the primary and secondary transitions of mouse pancreas development. We determined that the elimination of Neurod1 impairs the expression of key transcription factors for α- and β-cell differentiation, β-cell proliferation, insulin production, and islets of Langerhans formation. These findings demonstrate that the Neurod1 deletion altered the properties of α and β endocrine cells, resulting in severe neonatal diabetes, and thus, NEUROD1 is required for proper activation of the transcriptional network and differentiation of functional α and β cells.
    Keywords:  NEUROD1; genetic mutation; mouse model; pancreatic development; transcriptional network
    DOI:  https://doi.org/10.3390/ijms22136713
  15. Front Cell Dev Biol. 2021 ;9 682561
    Genome-Wide Histone H3K27 Acetylation Profiling Identified Genes Correlated With Prognosis in Papillary Thyroid Carcinoma.
    Luyao Zhang, Dan Xiong, Qian Liu, Yiling Luo, Yuhan Tian, Xi Xiao, Ye Sang, Yihao Liu, Shubin Hong, Shuang Yu, Jie Li, Weiming Lv, Yanbing Li, Zhonghui Tang, Rengyun Liu, Qian Zhong, Haipeng Xiao.
      Thyroid carcinoma (TC) is the most common endocrine malignancy, and papillary TC (PTC) is the most frequent subtype of TC, accounting for 85-90% of all the cases. Aberrant histone acetylation contributes to carcinogenesis by inducing the dysregulation of certain cancer-related genes. However, the histone acetylation landscape in PTC remains elusive. Here, we interrogated the epigenomes of PTC and benign thyroid nodule (BTN) tissues by applying H3K27ac chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) along with RNA-sequencing. By comparing the epigenomic features between PTC and BTN, we detected changes in H3K27ac levels at active regulatory regions, identified PTC-specific super-enhancer-associated genes involving immune-response and cancer-related pathways, and uncovered several genes that associated with disease-free survival of PTC. In summary, our data provided a genome-wide landscape of histone modification in PTC and demonstrated the role of enhancers in transcriptional regulations associated with prognosis of PTC.
    Keywords:  H3K27ac; benign thyroid nodule; enhancer; epigenetics; papillary thyroid carcinoma; transcriptome
    DOI:  https://doi.org/10.3389/fcell.2021.682561
  16. BMC Genomics. 2021 Jul 03. 22(1): 497
    A functional map of genomic HIF1α-DNA complexes in the eye lens revealed through multiomics analysis.
    Joshua Disatham, Lisa Brennan, Daniel Chauss, Jason Kantorow, Behdad Afzali, Marc Kantorow.
      BACKGROUND: During eye lens development the embryonic vasculature regresses leaving the lens without a direct oxygen source. Both embryonically and throughout adult life, the lens contains a decreasing oxygen gradient from the surface to the core that parallels the natural differentiation of immature surface epithelial cells into mature core transparent fiber cells. These properties of the lens suggest a potential role for hypoxia and the master regulator of the hypoxic response, hypoxia-inducible transcription factor 1 (HIF1), in the regulation of genes required for lens fiber cell differentiation, structure and transparency. Here, we employed a multiomics approach combining CUT&RUN, RNA-seq and ATACseq analysis to establish the genomic complement of lens HIF1α binding sites, genes activated or repressed by HIF1α and the chromatin states of HIF1α-regulated genes.RESULTS: CUT&RUN analysis revealed 8375 HIF1α-DNA binding complexes in the chick lens genome. One thousand one hundred ninety HIF1α-DNA binding complexes were significantly clustered within chromatin accessible regions (χ2 test p < 1 × 10- 55) identified by ATACseq. Formation of the identified HIF1α-DNA complexes paralleled the activation or repression of 526 genes, 116 of which contained HIF1α binding sites within 10kB of the transcription start sites. Some of the identified HIF1α genes have previously established lens functions while others have novel functions never before examined in the lens. GO and pathway analysis of these genes implicate HIF1α in the control of a wide-variety of cellular pathways potentially critical for lens fiber cell formation, structure and function including glycolysis, cell cycle regulation, chromatin remodeling, Notch and Wnt signaling, differentiation, development, and transparency.
    CONCLUSIONS: These data establish the first functional map of genomic HIF1α-DNA complexes in the eye lens. They identify HIF1α as an important regulator of a wide-variety of genes previously shown to be critical for lens formation and function and they reveal a requirement for HIF1α in the regulation of a wide-variety of genes not yet examined for lens function. They support a requirement for HIF1α in lens fiber cell formation, structure and function and they provide a basis for understanding the potential roles and requirements for HIF1α in the development, structure and function of more complex tissues.
    Keywords:  ATAC-seq; CUT&RUN; Chromatin; DNA binding; Gene regulation; HIF1α; Hypoxia; RNA-seq; Transcriptional regulation
    DOI:  https://doi.org/10.1186/s12864-021-07795-9
  17. Nucleic Acids Res. 2021 Jun 28. pii: gkab558. [Epub ahead of print]
    Cell-free genomics reveal intrinsic, cooperative and competitive determinants of chromatin interactions.
    Nikolas Eggers, Peter B Becker.
      Metazoan transcription factors distinguish their response elements from a large excess of similar sequences. We explored underlying principles of DNA shape read-out and factor cooperativity in chromatin using a unique experimental system. We reconstituted chromatin on Drosophila genomes in extracts of preblastoderm embryos, mimicking the naïve state of the zygotic genome prior to developmental transcription activation. We then compared the intrinsic binding specificities of three recombinant transcription factors, alone and in combination, with GA-rich recognition sequences genome-wide. For MSL2, all functional elements reside on the X chromosome, allowing to distinguish physiological elements from non-functional 'decoy' sites. The physiological binding profile of MSL2 is approximated through interaction with other factors: cooperativity with CLAMP and competition with GAF, which sculpts the profile by occluding non-functional sites. An extended DNA shape signature is differentially read out in chromatin. Our results reveal novel aspects of target selection in a complex chromatin environment.
    DOI:  https://doi.org/10.1093/nar/gkab558
  18. Nat Genet. 2021 Jul 01.
    Genetic drivers of m6A methylation in human brain, lung, heart and muscle.
    Xushen Xiong, Lei Hou, Yongjin P Park, Benoit Molinie, , Richard I Gregory, Manolis Kellis.
      The most prevalent post-transcriptional mRNA modification, N6-methyladenosine (m6A), plays diverse RNA-regulatory roles, but its genetic control in human tissues remains uncharted. Here we report 129 transcriptome-wide m6A profiles, covering 91 individuals and 4 tissues (brain, lung, muscle and heart) from GTEx/eGTEx. We integrate these with interindividual genetic and expression variation, revealing 8,843 tissue-specific and 469 tissue-shared m6A quantitative trait loci (QTLs), which are modestly enriched in, but mostly orthogonal to, expression QTLs. We integrate m6A QTLs with disease genetics, identifying 184 GWAS-colocalized m6A QTL, including brain m6A QTLs underlying neuroticism, depression, schizophrenia and anxiety; lung m6A QTLs underlying expiratory flow and asthma; and muscle/heart m6A QTLs underlying coronary artery disease. Last, we predict novel m6A regulators that show preferential binding in m6A QTLs, protein interactions with known m6A regulators and expression correlation with the m6A levels of their targets. Our results provide important insights and resources for understanding both cis and trans regulation of epitranscriptomic modifications, their interindividual variation and their roles in human disease.
    DOI:  https://doi.org/10.1038/s41588-021-00890-3
  19. Genome Res. 2021 Jun 30.
    A Bayesian inference transcription factor activity model for the analysis of single-cell transcriptomes.
    Shang Gao, Yang Dai, Jalees Rehman.
      Single-cell RNA sequencing (scRNA-seq) has emerged as a powerful experimental approach to study cellular heterogeneity. One of the challenges in scRNA-seq data analysis is integrating different types of biological data to consistently recognize discrete biological functions and regulatory mechanisms of cells, such as transcription factor activities and gene regulatory networks in distinct cell populations. We have developed an approach to infer transcription factor activities from scRNA-seq data that leverages existing biological data on transcription factor binding sites. The Bayesian inference transcription factor activity model (BITFAM) integrates ChIP-seq transcription factor binding information into scRNA-seq data analysis. We show that the inferred transcription factor activities for key cell types identify regulatory transcription factors that are known to mechanistically control cell function and cell fate. The BITFAM approach not only identifies biologically meaningful transcription factor activities, but also provides valuable insights into underlying transcription factor regulatory mechanisms.
    DOI:  https://doi.org/10.1101/gr.265595.120
  20. Cell Regen. 2021 Jul 02. 10(1): 17
    Forkhead box family transcription factors as versatile regulators for cellular reprogramming to pluripotency.
    Meijun Fu, Huan Chen, Zepo Cai, Yihang Yang, Ziyu Feng, Mengying Zeng, Lijun Chen, Yue Qin, Baomei Cai, Pinghui Zhu, Chunhua Zhou, Shengyong Yu, Jing Guo, Jing Liu, Shangtao Cao, Duanqing Pei.
      Forkhead box (Fox) transcription factors play important roles in mammalian development and disease. However, their function in mouse somatic cell reprogramming remains unclear. Here, we report that FoxD subfamily and FoxG1 accelerate induced pluripotent stem cells (iPSCs) generation from mouse fibroblasts as early as day4 while FoxA and FoxO subfamily impede this process obviously. More importantly, FoxD3, FoxD4 and FoxG1 can replace Oct4 respectively and generate iPSCs with germline transmission together with Sox2 and Klf4. On the contrary, FoxO6 almost totally blocks reprogramming through inhibiting cell proliferation, suppressing the expression of pluripotent genes and hindering the process of mesenchymal to epithelial transition (MET). Thus, our study uncovers unexpected roles of Fox transcription factors in reprogramming and offers new insights into cell fate transition.
    DOI:  https://doi.org/10.1186/s13619-021-00078-4
  21. Oncogene. 2021 Jul 02.
    Direct phosphorylation and stabilization of HIF-1α by PIM1 kinase drives angiogenesis in solid tumors.
    Andrea L Casillas, Shailender S Chauhan, Rachel K Toth, Alva G Sainz, Amber N Clements, Corbin C Jensen, Paul R Langlais, Cindy K Miranti, Anne E Cress, Noel A Warfel.
      Angiogenesis is essential for the sustained growth of solid tumors. Hypoxia-inducible factor 1 (HIF-1) is a master regulator of angiogenesis and constitutive activation of HIF-1 is frequently observed in human cancers. Therefore, understanding the mechanisms governing the activation of HIF-1 is critical for successful therapeutic targeting of tumor angiogenesis. Herein, we establish a new regulatory mechanism responsible for the constitutive activation of HIF-1α in cancer, irrespective of oxygen tension. PIM1 kinase directly phosphorylates HIF-1α at threonine 455, a previously uncharacterized site within its oxygen-dependent degradation domain. This phosphorylation event disrupts the ability of prolyl hydroxylases to bind and hydroxylate HIF-1α, interrupting its canonical degradation pathway and promoting constitutive transcription of HIF-1 target genes. Moreover, phosphorylation of the analogous site in HIF-2α (S435) stabilizes the protein through the same mechanism, indicating post-translational modification within the oxygen-dependent degradation domain as a mechanism of regulating the HIF-α subunits. In vitro and in vivo models demonstrate that expression of PIM1 is sufficient to stabilize HIF-1α and HIF-2α in normoxia and stimulate angiogenesis in a HIF-1-dependent manner. CRISPR mutants of HIF-1α (Thr455D) promoted increased tumor growth, proliferation, and angiogenesis. Moreover, HIF-1α-T455D xenograft tumors were refractory to the anti-angiogenic and cytotoxic effects of PIM inhibitors. These data identify a new signaling axis responsible for hypoxia-independent activation of HIF-1 and expand our understanding of the tumorigenic role of PIM1 in solid tumors.
    DOI:  https://doi.org/10.1038/s41388-021-01915-1
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