bims-crepig 2020-08-02 papers

bims-crepig

Biomed News

on Chromatin regulation and epigenetics in cell fate and cancer

Issue of 2020–08–02
33 papers selected by
Connor Rogerson, University of Cambridge, MRC Cancer Unit

Model-based analysis of chromatin interactions from dCas9-Based CAPTURE-3C-seq.
DNA methylation repels binding of hypoxia-inducible transcription factors to maintain tumor immunotolerance.
Cross-species analysis of enhancer logic using deep learning.
Expanded encyclopaedias of DNA elements in the human and mouse genomes.
Perspectives on ENCODE.
Cross-species chromatin interactions drive transcriptional rewiring in Epstein-Barr virus-positive gastric adenocarcinoma.
Epigenetic regulator function through mouse gastrulation.
CTCF orchestrates long-range cohesin-driven V(D)J recombinational scanning.
Analysis of Local Chromatin States Reveals Gene Transcription Potential during Mouse Neural Progenitor Cell Differentiation.
5-Formylcytosine landscapes of human preimplantation embryos at single-cell resolution.
Occupancy maps of 208 chromatin-associated proteins in one human cell type.
Landscape of cohesin-mediated chromatin loops in the human genome.
The changing mouse embryo transcriptome at whole tissue and single-cell resolution.
An integrative ENCODE resource for cancer genomics.
Interplay among ATP-Dependent Chromatin Remodelers Determines Chromatin Organisation in Yeast.
Spatiotemporal DNA methylome dynamics of the developing mouse fetus.
Regenerative Reprogramming of the Intestinal Stem Cell State via Hippo Signaling Suppresses Metastatic Colorectal Cancer.
NADP modulates RNA m6A methylation and adipogenesis via enhancing FTO activity.
Supervised enhancer prediction with epigenetic pattern recognition and targeted validation.
Epigenetic regulation of cancer stem cell and tumorigenesis.
Trib1 promotes acute myeloid leukemia progression by modulating the transcriptional programs of Hoxa9.
O-GlcNAcylation of Mef2c regulates myoblast differentiation.
Activation of NRF2 ameliorates oxidative stress and cystogenesis in autosomal dominant polycystic kidney disease.
A comprehensive evaluation of computational tools to identify differential methylation regions using RRBS data.
Epigenetic cell fate in Candida albicans is controlled by transcription factor condensates acting at super-enhancer-like elements.
An atlas of dynamic chromatin landscapes in mouse fetal development.
Mediator complex subunit Med19 binds directly GATA transcription factors and is required with Med1 for GATA-driven gene regulation in vivo.
Global reference mapping of human transcription factor footprints.
HDAC4 inhibition disrupts TET2 function in high-risk MDS and AML.
Landscape of DNA binding signatures of myocyte enhancer factor-2B reveals a unique interplay of base and shape readout.
Histone deacetylase 3 controls lung alveolar macrophage development and homeostasis.
Dissecting transcriptional amplification by MYC.
dream: Powerful differential expression analysis for repeated measures designs.

PLoS One. 2020 ;15(7): e0236666

Model-based analysis of chromatin interactions from dCas9-Based CAPTURE-3C-seq.

Yong Chen, Yunfei Wang, Xin Liu, Jian Xu, Michael Q Zhang.

Deciphering long-range chromatin interactions is critical for understanding temporal and tissue-specific gene expression regulated by cis- and trans-acting factors. By combining the chromosome conformation capture (3C) and biotinylated dCas9 system, we previously established a method CAPTURE-3C-seq to unbiasedly identify high-resolution and locus-specific long-range DNA interactions. Here we present the statistical model and a flexible pipeline, C3S, for analysing CAPTURE-3C-seq or similar experimental data from raw sequencing reads to significantly interacting chromatin loci. C3S provides all steps for data processing, quality control and result illustration. It can automatically define the bin size based on the binding peak of the dCas9-targeted regions. Furthermore, it supports the analysis of intra- and inter-chromosomal interactions for different mammalian cell types. We successfully applied C3S across multiple datasets in human K562 cells and mouse embryonic stem cells (mESC) for detecting known and new chromatin interactions at multiple scales. Integrative and topological analysis of the interacted loci at the human β-globin gene cluster provides new insights into mechanisms in developmental gene regulation and network structure in local chromosomal architecture. Furthermore, computational results in mESCs reveal a role for chromatin interacting loops between enhancers and promoters in regulating alternative transcripts of the pluripotency gene OCT4.

DOI: https://doi.org/10.1371/journal.pone.0236666
Genome Biol. 2020 Jul 27. 21(1): 182

DNA methylation repels binding of hypoxia-inducible transcription factors to maintain tumor immunotolerance.

Flora D'Anna, Laurien Van Dyck, Jieyi Xiong, Hui Zhao, Rebecca V Berrens, Junbin Qian, Pawel Bieniasz-Krzywiec, Vikas Chandra, Luc Schoonjans, Jason Matthews, Julie De Smedt, Liesbeth Minnoye, Ricardo Amorim, Sepideh Khorasanizadeh, Qian Yu, Liyun Zhao, Marie De Borre, Savvas N Savvides, M Celeste Simon, Peter Carmeliet, Wolf Reik, Fraydoon Rastinejad, Massimiliano Mazzone, Bernard Thienpont, Diether Lambrechts.

   BACKGROUND: Hypoxia is pervasive in cancer and other diseases. Cells sense and adapt to hypoxia by activating hypoxia-inducible transcription factors (HIFs), but it is still an outstanding question why cell types differ in their transcriptional response to hypoxia.
RESULTS: We report that HIFs fail to bind CpG dinucleotides that are methylated in their consensus binding sequence, both in in vitro biochemical binding assays and in vivo studies of differentially methylated isogenic cell lines. Based on in silico structural modeling, we show that 5-methylcytosine indeed causes steric hindrance in the HIF binding pocket. A model wherein cell-type-specific methylation landscapes, as laid down by the differential expression and binding of other transcription factors under normoxia, control cell-type-specific hypoxia responses is observed. We also discover ectopic HIF binding sites in repeat regions which are normally methylated. Genetic and pharmacological DNA demethylation, but also cancer-associated DNA hypomethylation, expose these binding sites, inducing HIF-dependent expression of cryptic transcripts. In line with such cryptic transcripts being more prone to cause double-stranded RNA and viral mimicry, we observe low DNA methylation and high cryptic transcript expression in tumors with high immune checkpoint expression, but not in tumors with low immune checkpoint expression, where they would compromise tumor immunotolerance. In a low-immunogenic tumor model, DNA demethylation upregulates cryptic transcript expression in a HIF-dependent manner, causing immune activation and reducing tumor growth.
CONCLUSIONS: Our data elucidate the mechanism underlying cell-type-specific responses to hypoxia and suggest DNA methylation and hypoxia to underlie tumor immunotolerance.

Keywords:  Cancer; Cryptic transcripts; DNA methylation; HIF; Hypoxia; Immunotherapy; Transcription factor binding

DOI:  https://doi.org/10.1186/s13059-020-02087-z
Genome Res. 2020 Jul 30. pii: gr.260844.120. [Epub ahead of print]

Cross-species analysis of enhancer logic using deep learning.

Liesbeth Minnoye, Ibrahim Ihsan Taskiran, David Mauduit, Maurizio Fazio, Linde Van Aerschot, Gert Hulselmans, Valerie Christiaens, Samira Makhzami, Monika Seltenhammer, Panagiotis Karras, Aline Primot, Edouard Cadieu, Ellen van Rooijen, Jean-Christophe Marine, Giorgia Egidy, Ghanem Elias Ghanem, Leonard Zon, Jasper Wouters, Stein Aerts.

Deciphering the genomic regulatory code of enhancers is a key challenge in biology as this code underlies cellular identity. A better understanding of how enhancers work will improve the interpretation of noncoding genome variation, and empower the generation of cell type-specific drivers for gene therapy. Here we explore the combination of deep learning and cross-species chromatin accessibility profiling to build explainable enhancer models. We apply this strategy to decipher the enhancer code in melanoma, a relevant case study due to the presence of distinct melanoma cell states. We trained and validated a deep learning model, called DeepMEL, using chromatin accessibility data of 26 melanoma samples across six different species. We demonstrate the accuracy of DeepMEL predictions on the CAGI5 challenge, where it significantly outperforms existing models on the melanoma enhancer of IRF4 Next, we exploit DeepMEL to analyse enhancer architectures and identify accurate transcription factor binding sites for the core regulatory complexes in the two different melanoma states, with distinct roles for each transcription factor, in terms of nucleosome displacement or enhancer activation. Finally, DeepMEL identifies orthologous enhancers across distantly related species where sequence alignment fails, and the model highlights specific nucleotide substitutions that underlie enhancer turnover. DeepMEL can be used from the Kipoi database to predict and optimise candidate enhancers, and to prioritise enhancer mutations. In addition, our computational strategy can be applied to other cancer or normal cell types.

DOI: https://doi.org/10.1101/gr.260844.120
Nature. 2020 Jul;583(7818): 699-710

Expanded encyclopaedias of DNA elements in the human and mouse genomes.

ENCODE Project Consortium

The human and mouse genomes contain instructions that specify RNAs and proteins and govern the timing, magnitude, and cellular context of their production. To better delineate these elements, phase III of the Encyclopedia of DNA Elements (ENCODE) Project has expanded analysis of the cell and tissue repertoires of RNA transcription, chromatin structure and modification, DNA methylation, chromatin looping, and occupancy by transcription factors and RNA-binding proteins. Here we summarize these efforts, which have produced 5,992 new experimental datasets, including systematic determinations across mouse fetal development. All data are available through the ENCODE data portal (https://www.encodeproject.org), including phase II ENCODE1 and Roadmap Epigenomics2 data. We have developed a registry of 926,535 human and 339,815 mouse candidate cis-regulatory elements, covering 7.9 and 3.4% of their respective genomes, by integrating selected datatypes associated with gene regulation, and constructed a web-based server (SCREEN; http://screen.encodeproject.org) to provide flexible, user-defined access to this resource. Collectively, the ENCODE data and registry provide an expansive resource for the scientific community to build a better understanding of the organization and function of the human and mouse genomes.

DOI: https://doi.org/10.1038/s41586-020-2493-4
Nature. 2020 Jul;583(7818): 693-698

Perspectives on ENCODE.

ENCODE Project Consortium

The Encylopedia of DNA Elements (ENCODE) Project launched in 2003 with the long-term goal of developing a comprehensive map of functional elements in the human genome. These included genes, biochemical regions associated with gene regulation (for example, transcription factor binding sites, open chromatin, and histone marks) and transcript isoforms. The marks serve as sites for candidate cis-regulatory elements (cCREs) that may serve functional roles in regulating gene expression1. The project has been extended to model organisms, particularly the mouse. In the third phase of ENCODE, nearly a million and more than 300,000 cCRE annotations have been generated for human and mouse, respectively, and these have provided a valuable resource for the scientific community.

DOI: https://doi.org/10.1038/s41586-020-2449-8
Nat Genet. 2020 Jul 27.

Cross-species chromatin interactions drive transcriptional rewiring in Epstein-Barr virus-positive gastric adenocarcinoma.

Atsushi Okabe, Kie Kyon Huang, Keisuke Matsusaka, Masaki Fukuyo, Manjie Xing, Xuewen Ong, Takayuki Hoshii, Genki Usui, Motoaki Seki, Yasunobu Mano, Bahityar Rahmutulla, Teru Kanda, Takayoshi Suzuki, Sun Young Rha, Tetsuo Ushiku, Masashi Fukayama, Patrick Tan, Atsushi Kaneda.

Epstein-Barr virus (EBV) is associated with several human malignancies including 8-10% of gastric cancers (GCs). Genome-wide analysis of 3D chromatin topologies across GC lines, primary tissue and normal gastric samples revealed chromatin domains specific to EBV-positive GC, exhibiting heterochromatin-to-euchromatin transitions and long-range human-viral interactions with non-integrated EBV episomes. EBV infection in vitro suffices to remodel chromatin topology and function at EBV-interacting host genomic loci, converting H3K9me3+ heterochromatin to H3K4me1+/H3K27ac+ bivalency and unleashing latent enhancers to engage and activate nearby GC-related genes (for example TGFBR2 and MZT1). Higher-order epigenotypes of EBV-positive GC thus signify a novel oncogenic paradigm whereby non-integrative viral genomes can directly alter host epigenetic landscapes ('enhancer infestation'), facilitating proto-oncogene activation and tumorigenesis.

DOI: https://doi.org/10.1038/s41588-020-0665-7
Nature. 2020 Jul 29.

Epigenetic regulator function through mouse gastrulation.

Stefanie Grosswendt, Helene Kretzmer, Zachary D Smith, Abhishek Sampath Kumar, Sara Hetzel, Lars Wittler, Sven Klages, Bernd Timmermann, Shankar Mukherji, Alexander Meissner.

During ontogeny, proliferating cells become restricted in their fate through the combined action of cell-type-specific transcription factors and ubiquitous epigenetic machinery, which recognizes universally available histone residues or nucleotides in a context-dependent manner1,2. The molecular functions of these regulators are generally well understood, but assigning direct developmental roles to them is hampered by complex mutant phenotypes that often emerge after gastrulation3,4. Single-cell RNA sequencing and analytical approaches have explored this highly conserved, dynamic period across numerous model organisms5-8, including mouse9-18. Here we advance these strategies using a combined zygotic perturbation and single-cell RNA-sequencing platform in which many mutant mouse embryos can be assayed simultaneously, recovering robust morphological and transcriptional information across a panel of ten essential regulators. Deeper analysis of central Polycomb repressive complex (PRC) 1 and 2 components indicates substantial cooperativity, but distinguishes a dominant role for PRC2 in restricting the germline. Moreover, PRC mutant phenotypes emerge after gross epigenetic and transcriptional changes within the initial conceptus prior to gastrulation. Our experimental framework may eventually lead to a fully quantitative view of how cellular diversity emerges using an identical genetic template and from a single totipotent cell.

DOI: https://doi.org/10.1038/s41586-020-2552-x
Nature. 2020 Jul 27.

CTCF orchestrates long-range cohesin-driven V(D)J recombinational scanning.

Zhaoqing Ba, Jiangman Lou, Adam Yongxin Ye, Hai-Qiang Dai, Edward W Dring, Sherry G Lin, Suvi Jain, Nia Kyritsis, Kyong-Rim Kieffer-Kwon, Rafael Casellas, Frederick W Alt.

RAG endonuclease initiates V(D)J recombination in progenitor (pro)-B cells1. Upon binding a recombination center (RC)-based JH, RAG scans upstream chromatin via loop extrusion, potentially mediated by cohesin, to locate Ds and assemble a DJH-based RC2. CTCF looping factor-bound elements (CBEs) within IGCR1 upstream of Ds impede RAG-scanning3-5; but their inactivation allows scanning to proximal VHs where additional CBEs activate rearrangement and impede scanning any further upstream5. Distal VH utilization is thought to involve diffusional RC access following large-scale Igh locus contraction6-8. Here, we test the potential of linear RAG-scanning to mediate distal VH usage in G1-arrested v-Abl-pro-B cell lines9, which undergo robust D-to-JH but little VH-to-DJH rearrangements, presumably due to lack of locus contraction2,5. Through an auxin-inducible approach10, we degrade the cohesin-component Rad2110-12 or CTCF12,13 in these G1-arrested lines. Rad21 degradation eliminated all V(D)J recombination and RAG-scanning-associated interactions, except RC-located DQ52-to-JH joining in which synapsis occurs by diffusion2. Remarkably, while CTCF degradation suppressed most CBE-based chromatin interactions, it promoted robust RC interactions with, and robust VH-to-DJH joining of, distal VHs, with patterns similar to those of "locus-contracted" primary pro-B cells. Thus, down-modulation of CTCF-bound scanning-impediment activity promotes cohesin-driven RAG-scanning across the 2.7Mb Igh locus.

DOI: https://doi.org/10.1038/s41586-020-2578-0
Cell Rep. 2020 Jul 28. pii: S2211-1247(20)30934-7. [Epub ahead of print]32(4): 107953

Analysis of Local Chromatin States Reveals Gene Transcription Potential during Mouse Neural Progenitor Cell Differentiation.

Juan Yu, Chaoyang Xiong, Baowen Zhuo, Zengqi Wen, Jie Shen, Cuifang Liu, Luyuan Chang, Kehui Wang, Min Wang, Chenyi Wu, Xudong Wu, Xueqin Xu, Haihe Ruan, Guohong Li.

  Chromatin dynamics play a critical role in cell fate determination and maintenance by regulating the expression of genes essential for development and differentiation. In mouse embryonic stem cells (mESCs), maintenance of pluripotency coincides with a poised chromatin state containing active and repressive histone modifications. However, the structural features of poised chromatin are largely uncharacterized. By adopting mild time-course MNase-seq with computational analysis, the low-compact chromatin in mESCs is featured in two groups: one in more open regions, corresponding to an active state, and the other enriched with bivalent histone modifications, considered the poised state. A parameter called the chromatin opening potential index (COPI) is also devised to quantify the transcription potential based on the dynamic changes of MNase-seq signals at promoter regions. Use of COPI provides effective prediction of gene activation potential and, more importantly, reveals a few developmental factors essential for mouse neural progenitor cell (NPC) differentiation.

Keywords:  MNase-seq; cell fate; poised chromatin; transcription potential

DOI:  https://doi.org/10.1016/j.celrep.2020.107953
PLoS Biol. 2020 Jul 30. 18(7): e3000799

5-Formylcytosine landscapes of human preimplantation embryos at single-cell resolution.

Yun Gao, Lin Li, Peng Yuan, Fan Zhai, Yixin Ren, Liying Yan, Rong Li, Ying Lian, Xiaohui Zhu, Xinglong Wu, Kehkooi Kee, Lu Wen, Jie Qiao, Fuchou Tang.

Epigenetic dynamics, such as DNA methylation and chromatin accessibility, have been extensively explored in human preimplantation embryos. However, the active demethylation process during this crucial period remains largely unexplored. In this study, we use single-cell chemical-labeling-enabled C-to-T conversion sequencing (CLEVER-seq) to quantify the DNA 5-formylcytosine (5fC) levels of human preimplantation embryos. We find that 5-formylcytosine phosphate guanine (5fCpG) exhibits genomic element-specific distribution features and is enriched in L1 and endogenous retrovirus-K (ERVK), the subfamilies of repeat elements long interspersed nuclear elements (LINEs) and long terminal repeats (LTRs), respectively. Unlike in mice, paired pronuclei in the same zygote present variable difference of 5fCpG levels, although the male pronuclei experience stronger global demethylation. The nucleosome-occupied regions show a higher 5fCpG level compared with nucleosome-depleted ones, suggesting the role of 5fC in organizing nucleosome position. Collectively, our work offers a valuable resource for ten-eleven translocation protein family (TET)-dependent active demethylation-related study during human early embryonic development.

DOI: https://doi.org/10.1371/journal.pbio.3000799
Nature. 2020 Jul;583(7818): 720-728

Occupancy maps of 208 chromatin-associated proteins in one human cell type.

E Christopher Partridge, Surya B Chhetri, Jeremy W Prokop, Ryne C Ramaker, Camden S Jansen, Say-Tar Goh, Mark Mackiewicz, Kimberly M Newberry, Laurel A Brandsmeier, Sarah K Meadows, C Luke Messer, Andrew A Hardigan, Candice J Coppola, Emma C Dean, Shan Jiang, Daniel Savic, Ali Mortazavi, Barbara J Wold, Richard M Myers, Eric M Mendenhall.

Transcription factors are DNA-binding proteins that have key roles in gene regulation1,2. Genome-wide occupancy maps of transcriptional regulators are important for understanding gene regulation and its effects on diverse biological processes3-6. However, only a minority of the more than 1,600 transcription factors encoded in the human genome has been assayed. Here we present, as part of the ENCODE (Encyclopedia of DNA Elements) project, data and analyses from chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) experiments using the human HepG2 cell line for 208 chromatin-associated proteins (CAPs). These comprise 171 transcription factors and 37 transcriptional cofactors and chromatin regulator proteins, and represent nearly one-quarter of CAPs expressed in HepG2 cells. The binding profiles of these CAPs form major groups associated predominantly with promoters or enhancers, or with both. We confirm and expand the current catalogue of DNA sequence motifs for transcription factors, and describe motifs that correspond to other transcription factors that are co-enriched with the primary ChIP target. For example, FOX family motifs are enriched in ChIP-seq peaks of 37 other CAPs. We show that motif content and occupancy patterns can distinguish between promoters and enhancers. This catalogue reveals high-occupancy target regions at which many CAPs associate, although each contains motifs for only a minority of the numerous associated transcription factors. These analyses provide a more complete overview of the gene regulatory networks that define this cell type, and demonstrate the usefulness of the large-scale production efforts of the ENCODE Consortium.

DOI: https://doi.org/10.1038/s41586-020-2023-4
Nature. 2020 Jul;583(7818): 737-743

Landscape of cohesin-mediated chromatin loops in the human genome.

Fabian Grubert, Rohith Srivas, Damek V Spacek, Maya Kasowski, Mariana Ruiz-Velasco, Nasa Sinnott-Armstrong, Peyton Greenside, Anil Narasimha, Qing Liu, Benjamin Geller, Akshay Sanghi, Michael Kulik, Silin Sa, Marlene Rabinovitch, Anshul Kundaje, Stephen Dalton, Judith B Zaugg, Michael Snyder.

Physical interactions between distal regulatory elements have a key role in regulating gene expression, but the extent to which these interactions vary between cell types and contribute to cell-type-specific gene expression remains unclear. Here, to address these questions as part of phase III of the Encyclopedia of DNA Elements (ENCODE), we mapped cohesin-mediated chromatin loops, using chromatin interaction analysis by paired-end tag sequencing (ChIA-PET), and analysed gene expression in 24 diverse human cell types, including core ENCODE cell lines. Twenty-eight per cent of all chromatin loops vary across cell types; these variations modestly correlate with changes in gene expression and are effective at grouping cell types according to their tissue of origin. The connectivity of genes corresponds to different functional classes, with housekeeping genes having few contacts, and dosage-sensitive genes being more connected to enhancer elements. This atlas of chromatin loops complements the diverse maps of regulatory architecture that comprise the ENCODE Encyclopedia, and will help to support emerging analyses of genome structure and function.

DOI: https://doi.org/10.1038/s41586-020-2151-x
Nature. 2020 Jul;583(7818): 760-767

The changing mouse embryo transcriptome at whole tissue and single-cell resolution.

Peng He, Brian A Williams, Diane Trout, Georgi K Marinov, Henry Amrhein, Libera Berghella, Say-Tar Goh, Ingrid Plajzer-Frick, Veena Afzal, Len A Pennacchio, Diane E Dickel, Axel Visel, Bing Ren, Ross C Hardison, Yu Zhang, Barbara J Wold.

During mammalian embryogenesis, differential gene expression gradually builds the identity and complexity of each tissue and organ system1. Here we systematically quantified mouse polyA-RNA from day 10.5 of embryonic development to birth, sampling 17 tissues and organs. The resulting developmental transcriptome is globally structured by dynamic cytodifferentiation, body-axis and cell-proliferation gene sets that were further characterized by the transcription factor motif codes of their promoters. We decomposed the tissue-level transcriptome using single-cell RNA-seq (sequencing of RNA reverse transcribed into cDNA) and found that neurogenesis and haematopoiesis dominate at both the gene and cellular levels, jointly accounting for one-third of differential gene expression and more than 40% of identified cell types. By integrating promoter sequence motifs with companion ENCODE epigenomic profiles, we identified a prominent promoter de-repression mechanism in neuronal expression clusters that was attributable to known and novel repressors. Focusing on the developing limb, single-cell RNA data identified 25 candidate cell types that included progenitor and differentiating states with computationally inferred lineage relationships. We extracted cell-type transcription factor networks and complementary sets of candidate enhancer elements by using single-cell RNA-seq to decompose integrative cis-element (IDEAS) models that were derived from whole-tissue epigenome chromatin data. These ENCODE reference data, computed network components and IDEAS chromatin segmentations are companion resources to the matching epigenomic developmental matrix, and are available for researchers to further mine and integrate.

DOI: https://doi.org/10.1038/s41586-020-2536-x
Nat Commun. 2020 Jul 29. 11(1): 3696

An integrative ENCODE resource for cancer genomics.

Jing Zhang, Donghoon Lee, Vineet Dhiman, Peng Jiang, Jie Xu, Patrick McGillivray, Hongbo Yang, Jason Liu, William Meyerson, Declan Clarke, Mengting Gu, Shantao Li, Shaoke Lou, Jinrui Xu, Lucas Lochovsky, Matthew Ung, Lijia Ma, Shan Yu, Qin Cao, Arif Harmanci, Koon-Kiu Yan, Anurag Sethi, Gamze Gürsoy, Michael Rutenberg Schoenberg, Joel Rozowsky, Jonathan Warrell, Prashant Emani, Yucheng T Yang, Timur Galeev, Xiangmeng Kong, Shuang Liu, Xiaotong Li, Jayanth Krishnan, Yanlin Feng, Juan Carlos Rivera-Mulia, Jessica Adrian, James R Broach, Michael Bolt, Jennifer Moran, Dominic Fitzgerald, Vishnu Dileep, Tingting Liu, Shenglin Mei, Takayo Sasaki, Claudia Trevilla-Garcia, Su Wang, Yanli Wang, Chongzhi Zang, Daifeng Wang, Robert J Klein, Michael Snyder, David M Gilbert, Kevin Yip, Chao Cheng, Feng Yue, X Shirley Liu, Kevin P White, Mark Gerstein.

ENCODE comprises thousands of functional genomics datasets, and the encyclopedia covers hundreds of cell types, providing a universal annotation for genome interpretation. However, for particular applications, it may be advantageous to use a customized annotation. Here, we develop such a custom annotation by leveraging advanced assays, such as eCLIP, Hi-C, and whole-genome STARR-seq on a number of data-rich ENCODE cell types. A key aspect of this annotation is comprehensive and experimentally derived networks of both transcription factors and RNA-binding proteins (TFs and RBPs). Cancer, a disease of system-wide dysregulation, is an ideal application for such a network-based annotation. Specifically, for cancer-associated cell types, we put regulators into hierarchies and measure their network change (rewiring) during oncogenesis. We also extensively survey TF-RBP crosstalk, highlighting how SUB1, a previously uncharacterized RBP, drives aberrant tumor expression and amplifies the effect of MYC, a well-known oncogenic TF. Furthermore, we show how our annotation allows us to place oncogenic transformations in the context of a broad cell space; here, many normal-to-tumor transitions move towards a stem-like state, while oncogene knockdowns show an opposing trend. Finally, we organize the resource into a coherent workflow to prioritize key elements and variants, in addition to regulators. We showcase the application of this prioritization to somatic burdening, cancer differential expression and GWAS. Targeted validations of the prioritized regulators, elements and variants using siRNA knockdowns, CRISPR-based editing, and luciferase assays demonstrate the value of the ENCODE resource.

DOI: https://doi.org/10.1038/s41467-020-14743-w
Biology (Basel). 2020 Jul 25. pii: E190. [Epub ahead of print]9(8):

Interplay among ATP-Dependent Chromatin Remodelers Determines Chromatin Organisation in Yeast.

Hemant K Prajapati, Josefina Ocampo, David J Clark.

  Cellular DNA is packaged into chromatin, which is composed of regularly-spaced nucleosomes with occasional gaps corresponding to active regulatory elements, such as promoters and enhancers, called nucleosome-depleted regions (NDRs). This chromatin organisation is primarily determined by the activities of a set of ATP-dependent remodeling enzymes that are capable of moving nucleosomes along DNA, or of evicting nucleosomes altogether. In yeast, the nucleosome-spacing enzymes are ISW1 (Imitation SWitch protein 1), Chromodomain-Helicase-DNA-binding (CHD)1, ISW2 (Imitation SWitch protein 2) and INOsitol-requiring 80 (INO80); the nucleosome eviction enzymes are the SWItching/Sucrose Non-Fermenting (SWI/SNF) family, the Remodeling the Structure of Chromatin (RSC) complexes and INO80. We discuss the contributions of each set of enzymes to chromatin organisation. ISW1 and CHD1 are the major spacing enzymes; loss of both enzymes results in major chromatin disruption, partly due to the appearance of close-packed di-nucleosomes. ISW1 and CHD1 compete to set nucleosome spacing on most genes. ISW1 is dominant, setting wild type spacing, whereas CHD1 sets short spacing and may dominate on highly-transcribed genes. We propose that the competing remodelers regulate spacing, which in turn controls the binding of linker histone (H1) and therefore the degree of chromatin folding. Thus, genes with long spacing bind more H1, resulting in increased chromatin compaction. RSC, SWI/SNF and INO80 are involved in NDR formation, either directly by nucleosome eviction or repositioning, or indirectly by affecting the size of the complex that resides in the NDR. The nature of this complex is controversial: some suggest that it is a RSC-bound "fragile nucleosome", whereas we propose that it is a non-histone transcription complex. In either case, this complex appears to serve as a barrier to nucleosome formation, resulting in the formation of phased nucleosomal arrays on both sides.

Keywords:  CHD1; Chromatin; Chromatin remodelers; INO80; ISW1; ISW2; RSC; SWI/SNF; nucleosome phasing; nucleosome spacing

DOI:  https://doi.org/10.3390/biology9080190
Nature. 2020 Jul;583(7818): 752-759

Spatiotemporal DNA methylome dynamics of the developing mouse fetus.

Yupeng He, Manoj Hariharan, David U Gorkin, Diane E Dickel, Chongyuan Luo, Rosa G Castanon, Joseph R Nery, Ah Young Lee, Yuan Zhao, Hui Huang, Brian A Williams, Diane Trout, Henry Amrhein, Rongxin Fang, Huaming Chen, Bin Li, Axel Visel, Len A Pennacchio, Bing Ren, Joseph R Ecker.

Cytosine DNA methylation is essential for mammalian development but understanding of its spatiotemporal distribution in the developing embryo remains limited1,2. Here, as part of the mouse Encyclopedia of DNA Elements (ENCODE) project, we profiled 168 methylomes from 12 mouse tissues or organs at 9 developmental stages from embryogenesis to adulthood. We identified 1,808,810 genomic regions that showed variations in CG methylation by comparing the methylomes of different tissues or organs from different developmental stages. These DNA elements predominantly lose CG methylation during fetal development, whereas the trend is reversed after birth. During late stages of fetal development, non-CG methylation accumulated within the bodies of key developmental transcription factor genes, coinciding with their transcriptional repression. Integration of genome-wide DNA methylation, histone modification and chromatin accessibility data enabled us to predict 461,141 putative developmental tissue-specific enhancers, the human orthologues of which were enriched for disease-associated genetic variants. These spatiotemporal epigenome maps provide a resource for studies of gene regulation during tissue or organ progression, and a starting point for investigating regulatory elements that are involved in human developmental disorders.

DOI: https://doi.org/10.1038/s41586-020-2119-x
Cell Stem Cell. 2020 Jul 24. pii: S1934-5909(20)30341-6. [Epub ahead of print]

Regenerative Reprogramming of the Intestinal Stem Cell State via Hippo Signaling Suppresses Metastatic Colorectal Cancer.

Priscilla Cheung, Jordi Xiol, Michael T Dill, Wei-Chien Yuan, Riccardo Panero, Jatin Roper, Fernando G Osorio, Dejan Maglic, Qi Li, Basanta Gurung, Raffaele A Calogero, Ömer H Yilmaz, Junhao Mao, Fernando D Camargo.

  Although the Hippo transcriptional coactivator YAP is considered oncogenic in many tissues, its roles in intestinal homeostasis and colorectal cancer (CRC) remain controversial. Here, we demonstrate that the Hippo kinases LATS1/2 and MST1/2, which inhibit YAP activity, are required for maintaining Wnt signaling and canonical stem cell function. Hippo inhibition induces a distinct epithelial cell state marked by low Wnt signaling, a wound-healing response, and transcription factor Klf6 expression. Notably, loss of LATS1/2 or overexpression of YAP is sufficient to reprogram Lgr5+ cancer stem cells to this state and thereby suppress tumor growth in organoids, patient-derived xenografts, and mouse models of primary and metastatic CRC. Finally, we demonstrate that genetic deletion of YAP and its paralog TAZ promotes the growth of these tumors. Collectively, our results establish the role of YAP as a tumor suppressor in the adult colon and implicate Hippo kinases as therapeutic vulnerabilities in colorectal malignancies.

Keywords:  Hippo signaling; Wnt signaling; colorectal cancer; intestinal stem cells; metastasis; regeneration

DOI:  https://doi.org/10.1016/j.stem.2020.07.003
Nat Chem Biol. 2020 Jul 27.

NADP modulates RNA m6A methylation and adipogenesis via enhancing FTO activity.

Lina Wang, Chengli Song, Na Wang, Songyu Li, Qiaoling Liu, Zhen Sun, Kai Wang, Shi-Cang Yu, Qingkai Yang.

Metabolism is often regulated by the transcription and translation of RNA. In turn, it is likely that some metabolites regulate enzymes controlling reversible RNA modification, such as N6-methyladenosine (m6A), to modulate RNA. This hypothesis is at least partially supported by the findings that multiple metabolic diseases are highly associated with fat mass and obesity-associated protein (FTO), an m6A demethylase. However, knowledge about whether and which metabolites directly regulate m6A remains elusive. Here, we show that NADP directly binds FTO, independently increases FTO activity, and promotes RNA m6A demethylation and adipogenesis. We screened a set of metabolites using a fluorescence quenching assay and NADP was identified to remarkably bind FTO. In vitro demethylation assays indicated that NADP enhances FTO activity. Furthermore, NADP regulated mRNA m6A via FTO in vivo, and deletion of FTO blocked NADP-enhanced adipogenesis in 3T3-L1 preadipocytes. These results build a direct link between metabolism and RNA m6A demethylation.

DOI: https://doi.org/10.1038/s41589-020-0601-2
Nat Methods. 2020 Aug;17(8): 807-814

Supervised enhancer prediction with epigenetic pattern recognition and targeted validation.

Anurag Sethi, Mengting Gu, Emrah Gumusgoz, Landon Chan, Koon-Kiu Yan, Joel Rozowsky, Iros Barozzi, Veena Afzal, Jennifer A Akiyama, Ingrid Plajzer-Frick, Chengfei Yan, Catherine S Novak, Momoe Kato, Tyler H Garvin, Quan Pham, Anne Harrington, Brandon J Mannion, Elizabeth A Lee, Yoko Fukuda-Yuzawa, Axel Visel, Diane E Dickel, Kevin Y Yip, Richard Sutton, Len A Pennacchio, Mark Gerstein.

Enhancers are important non-coding elements, but they have traditionally been hard to characterize experimentally. The development of massively parallel assays allows the characterization of large numbers of enhancers for the first time. Here, we developed a framework using Drosophila STARR-seq to create shape-matching filters based on meta-profiles of epigenetic features. We integrated these features with supervised machine-learning algorithms to predict enhancers. We further demonstrated that our model could be transferred to predict enhancers in mammals. We comprehensively validated the predictions using a combination of in vivo and in vitro approaches, involving transgenic assays in mice and transduction-based reporter assays in human cell lines (153 enhancers in total). The results confirmed that our model can accurately predict enhancers in different species without re-parameterization. Finally, we examined the transcription factor binding patterns at predicted enhancers versus promoters. We demonstrated that these patterns enable the construction of a secondary model that effectively distinguishes enhancers and promoters.

DOI: https://doi.org/10.1038/s41592-020-0907-8
Adv Cancer Res. 2020 ;pii: S0065-230X(20)30043-9. [Epub ahead of print]148 1-26

Epigenetic regulation of cancer stem cell and tumorigenesis.

Kezhou Zhu, Victoria Xie, Suyun Huang.

  As a unique subpopulation of cancer cells, cancer stem cells (CSCs) acquire the resistance to conventional therapies and appear to be the prime cause of cancer recurrence. Like their normal counterparts, CSCs can renew themselves and generate differentiated progenies. Cancer stem cells are distinguished among heterogenous cancer cells by molecular markers and their capacity of efficiently forming new tumors composed of diverse and heterogenous cancer cells. Tumor heterogeneity can be inter- or intra-tumor, molecularly resulting from the accumulation of genetic and non-genetic alterations. Non-genetic alterations are mainly changes on epigenetic modifications of DNA and histone, and chromatin remodeling. As tumor-initiating cells and contributing to the tumor heterogeneity in the brain, glioblastoma stem cells (GSCs) attract extensive research interests. Epigenetic modifications confer on tumor cells including CSCs reversible and inheritable genomic changes and affect gene expression without alteration in DNA sequence. Here, we will review recent advances in histone demethylation, DNA methylation, RNA methylation and ubiquitination in glioblastomas and their impacts on tumorigenesis with a focus on CSCs.

Keywords:  Epigenetic regulation; Gene expression; Glioblastoma stem cell; Tumorigenesis

DOI:  https://doi.org/10.1016/bs.acr.2020.05.001
Blood. 2020 Jul 30. pii: blood.2019004586. [Epub ahead of print]

Trib1 promotes acute myeloid leukemia progression by modulating the transcriptional programs of Hoxa9.

Seiko Yoshino, Takashi Yokoyama, Yoshitaka Sunami, Tomoko Takahara, Aya Nakamura, Yukari Yamazaki, Shuichi Tsutsumi, Hiroyuki Aburatani, Takuro Nakamura.

The pseudokinase Trib1 functions as a myeloid oncogene that recruits the E3 ubiquitin ligase COP1 to C/EBPa and interacts with MEK1 to enhance ERK phosphorylation. Close genetic effect of Trib1 on Hoxa9 has been observed in myeloid leukemogenesis where Trib1 overexpression significantly accelerates Hoxa9-induced leukemia onset. However, the mechanism underlying how Trib1 functionally modulates Hoxa9 transcription activity is unclear. Herein, we provide evidence that Trib1 modulates Hoxa9-associated super-enhancers. ChIP-seq analysis identified increased histone H3K27Ac signals at super-enhancers of the Erg, Spns2, Rgl1, and Pik3cd loci, as well as increased mRNA expression of these genes. Modification of super-enhancer activity was mostly achieved via the degradation of C/EBPa p42 by Trib1, with a slight contribution from the MEK/ERK pathway. Silencing of Erg abrogated the growth advantage acquired by Trib1 overexpression, indicating that Erg is a critical downstream target of the Trib1/Hoxa9 axis. Moreover, treatment of acute myeloid leukemia (AML) cells with the BRD4 inhibitor JQ1 showed growth inhibition in a Trib1/Erg-dependent manner both in vitro and in vivo. Upregulation of ERG by TRIB1 was also observed in human AML cell lines, suggesting that Trib1 is a potential therapeutic target of Hoxa9-associated AML. Taken together, our study demonstrates a novel mechanism by which Trib1 modulates chromatin and Hoxa9-driven transcription in myeloid leukemogenesis.

DOI: https://doi.org/10.1182/blood.2019004586
Biochem Biophys Res Commun. 2020 Aug 27. pii: S0006-291X(20)31232-8. [Epub ahead of print]529(3): 692-698

O-GlcNAcylation of Mef2c regulates myoblast differentiation.

Han Byeol Kim, Hyeon Gyu Seo, SeongJin Son, Hyeonjin Choi, Byung Gyu Kim, Tae Hyun Kweon, Sunghoon Kim, Jaeyoung Pai, Injae Shin, Won Ho Yang, Jin Won Cho.

  Unlike other types of glycosylation, O-GlcNAcylation is a single glycosylation which occurs exclusively in the nucleus and cytosol. O-GlcNAcylation underlie metabolic diseases, including diabetes and obesity. Furthermore, O-GlcNAcylation affects different oncogenic processes such as osteoblast differentiation, adipogenesis and hematopoiesis. Emerging evidence suggests that skeletal muscle differentiation is also regulated by O-GlcNAcylation, but the detailed molecular mechanism has not been fully elucidated. In this study, we showed that hyper-O-GlcNAcylation reduced the expression of myogenin, a transcription factor critical for terminal muscle development, in C2C12 myoblasts differentiation by O-GlcNAcylation on Thr9 of myocyte-specific enhancer factor 2c. Furthermore, we showed that O-GlcNAcylation on Mef2c inhibited its DNA binding affinity to myogenin promoter. Taken together, we demonstrated that hyper-O-GlcNAcylation attenuates skeletal muscle differentiation by increased O-GlcNAcylation on Mef2c, which downregulates its DNA binding affinity.

Keywords:  Mef2c; Myoblast differentiation; Myogenin; O-GlcNAc

DOI:  https://doi.org/10.1016/j.bbrc.2020.06.031
Sci Transl Med. 2020 Jul 29. pii: eaba3613. [Epub ahead of print]12(554):

Activation of NRF2 ameliorates oxidative stress and cystogenesis in autosomal dominant polycystic kidney disease.

Yi Lu, Yongzhan Sun, Zhiheng Liu, Yumei Lu, Xu Zhu, Bingxue Lan, Zeyun Mi, Lin Dang, Na Li, Wenlei Zhan, Lu Tan, Jingbo Pi, Hui Xiong, Lirong Zhang, Yupeng Chen.

Oxidative stress is emerging as a crucial contributor to the pathogenesis of autosomal dominant polycystic kidney disease (ADPKD), but the molecular mechanisms underlying the disturbed redox homeostasis in cystic cells remain elusive. Here, we identified the impaired activity of the NRF2 (nuclear factor erythroid 2-related factor 2) antioxidant pathway as a driver of oxidative damage and ADPKD progression. Using a quantitative proteomic approach, together with biochemical analyses, we found that increased degradation of NRF2 protein suppressed the NRF2 antioxidant pathway in ADPKD mouse kidneys. In a cohort of patients with ADPKD, reactive oxygen species (ROS) frequently accumulated, and their production correlated negatively with NRF2 abundance and positively with disease severity. In an orthologous ADPKD mouse model, genetic deletion of Nrf2 further increased ROS generation and promoted cyst growth, whereas pharmacological induction of NRF2 reduced ROS production and slowed cystogenesis and disease progression. Mechanistically, pharmacological induction of NRF2 remodeled enhancer landscapes and activated NRF2-bound enhancer-associated genes in ADPKD cells. The activation domain of NRF2 formed phase-separated condensates with MEDIATOR complex subunit MED16 in vitro, and optimal Mediator recruitment to genomic loci depended on NRF2 in vivo. Together, these findings indicate that NRF2 remodels enhancer landscapes and activates its target genes through a phase separation mechanism and that activation of NRF2 represents a promising strategy for restoring redox homeostasis and combatting ADPKD.

DOI: https://doi.org/10.1126/scitranslmed.aba3613
Genomics. 2020 Jul 23. pii: S0888-7543(20)31059-4. [Epub ahead of print]

A comprehensive evaluation of computational tools to identify differential methylation regions using RRBS data.

Yi Liu, Yi Han, Liyuan Zhou, Xiaoqing Pan, Xiwei Sun, Yong Liu, Mingyu Liang, Jiale Qin, Pengyuan Liu, Yan Lu.

  DNA methylation plays a vital role in transcription regulation. Reduced representation bisulfite sequencing (RRBS) is becoming common for analyzing genome-wide methylation profiles at the single nucleotide level. A major goal of RRBS studies is to detect differentially methylated regions (DMRs) between different biological conditions. The previous tools to predict DMRs lack consistency. Here, we simulated RRBS datasets with significant attributes of real sequencing data under a wide range of scenarios, and systematically evaluated seven DMR detection tools in terms of type I error rate, precision/recall (PR), and area under ROC curve (AUC) using different methylation levels, sequencing coverage depth, length of DMRs, read length, and sample sizes. DMRfinder, methylSig, and methylKit were our preferred tools for RRBS data analysis, in terms of their AUC and PR curves. Our comparison highlights the different applicability of DMR detection tools and provides information to guide researchers towards the advancement of sequence-based DMR analysis.

Keywords:  DNA methylation; Differentially methylated regions; Reduced representation bisulfite sequencing,

DOI:  https://doi.org/10.1016/j.ygeno.2020.07.032
Nat Microbiol. 2020 Jul 27.

Epigenetic cell fate in Candida albicans is controlled by transcription factor condensates acting at super-enhancer-like elements.

Corey Frazer, Mae I Staples, Yoori Kim, Matthew Hirakawa, Maureen A Dowell, Nicole V Johnson, Aaron D Hernday, Veronica H Ryan, Nicolas L Fawzi, Ilya J Finkelstein, Richard J Bennett.

Cell identity in eukaryotes is controlled by transcriptional regulatory networks that define cell-type-specific gene expression. In the opportunistic fungal pathogen Candida albicans, transcriptional regulatory networks regulate epigenetic switching between two alternative cell states, 'white' and 'opaque', that exhibit distinct host interactions. In the present study, we reveal that the transcription factors (TFs) regulating cell identity contain prion-like domains (PrLDs) that enable liquid-liquid demixing and the formation of phase-separated condensates. Multiple white-opaque TFs can co-assemble into complex condensates as observed on single DNA molecules. Moreover, heterotypic interactions between PrLDs support the assembly of multifactorial condensates at a synthetic locus within live eukaryotic cells. Mutation of the Wor1 TF revealed that substitution of acidic residues in the PrLD blocked its ability to phase separate and co-recruit other TFs in live cells, as well as its function in C. albicans cell fate determination. Together, these studies reveal that PrLDs support the assembly of TF complexes that control fungal cell identity and highlight parallels with the 'super-enhancers' that regulate mammalian cell fate.

DOI: https://doi.org/10.1038/s41564-020-0760-7
Nature. 2020 Jul;583(7818): 744-751

An atlas of dynamic chromatin landscapes in mouse fetal development.

David U Gorkin, Iros Barozzi, Yuan Zhao, Yanxiao Zhang, Hui Huang, Ah Young Lee, Bin Li, Joshua Chiou, Andre Wildberg, Bo Ding, Bo Zhang, Mengchi Wang, J Seth Strattan, Jean M Davidson, Yunjiang Qiu, Veena Afzal, Jennifer A Akiyama, Ingrid Plajzer-Frick, Catherine S Novak, Momoe Kato, Tyler H Garvin, Quan T Pham, Anne N Harrington, Brandon J Mannion, Elizabeth A Lee, Yoko Fukuda-Yuzawa, Yupeng He, Sebastian Preissl, Sora Chee, Jee Yun Han, Brian A Williams, Diane Trout, Henry Amrhein, Hongbo Yang, J Michael Cherry, Wei Wang, Kyle Gaulton, Joseph R Ecker, Yin Shen, Diane E Dickel, Axel Visel, Len A Pennacchio, Bing Ren.

The Encyclopedia of DNA Elements (ENCODE) project has established a genomic resource for mammalian development, profiling a diverse panel of mouse tissues at 8 developmental stages from 10.5 days after conception until birth, including transcriptomes, methylomes and chromatin states. Here we systematically examined the state and accessibility of chromatin in the developing mouse fetus. In total we performed 1,128 chromatin immunoprecipitation with sequencing (ChIP-seq) assays for histone modifications and 132 assay for transposase-accessible chromatin using sequencing (ATAC-seq) assays for chromatin accessibility across 72 distinct tissue-stages. We used integrative analysis to develop a unified set of chromatin state annotations, infer the identities of dynamic enhancers and key transcriptional regulators, and characterize the relationship between chromatin state and accessibility during developmental gene regulation. We also leveraged these data to link enhancers to putative target genes and demonstrate tissue-specific enrichments of sequence variants associated with disease in humans. The mouse ENCODE data sets provide a compendium of resources for biomedical researchers and achieve, to our knowledge, the most comprehensive view of chromatin dynamics during mammalian fetal development to date.

DOI: https://doi.org/10.1038/s41586-020-2093-3
J Biol Chem. 2020 Jul 31. pii: jbc.RA120.013728. [Epub ahead of print]

Mediator complex subunit Med19 binds directly GATA transcription factors and is required with Med1 for GATA-driven gene regulation in vivo.

Clément Immarigeon, Sandra Bernat-Fabre, Emmanuelle Guillou, Alexis Verger, Elodie Prince, Mohamed A Benmedjahed, Adeline Payet, Marie Couralet, Didier Monte, Vincent Villeret, Henri-Marc Bourbon, Muriel Boube.

  The evolutionarily conserved multiprotein Mediator complex (MED) serves as an interface between DNA-bound transcription factors (TFs) and the RNA Pol II machinery. It has been proposed that each TF interacts with a dedicated MED subunit to induce specific transcriptional responses. But are these binary partnerships sufficient to mediate TF functions? We have previously established that the Med1 Mediator subunit serves as a cofactor of GATA TFs in Drosophila, as shown in mammals. Here, we observe mutant phenotype similarities between another subunit, Med19, and the Drosophila GATA TF Pannier (Pnr), suggesting functional interaction. We further show that Med19 physically interacts with the Drosophila GATA TFs, Pnr and Serpent (Srp), in vivo and in vitro through their conserved C-zinc finger domains. Moreover, Med19 loss of function experiments in vivo or in cellulo indicate that it is required for Pnr- and Srp- dependent gene expression, suggesting general GATA cofactor functions. Interestingly, Med19 but not Med1 is critical for the regulation of all tested GATA target genes, implying shared or differential use of MED subunits by GATAs depending on the target gene. Lastly, we show a direct interaction between Med19 and Med1 by GST-pull-down experiments indicating privileged contacts between these two subunits of the MED middle module. Together, these findings identify Med19/Med1 as a composite GATA TF interface and suggest that binary MED subunit - TF partnerships are probably oversimplified models. We propose several mechanisms to account for the transcriptional regulation of GATAs-targeted genes.

Keywords:  GATA Pannier Pnr; GATA Serpent Srp; GATA transcription factor; Med19 Med1; Mediator complex MED; Thoracic closure; drosophila genetics; gene regulation; transcription coregulator; transcription regulation

DOI:  https://doi.org/10.1074/jbc.RA120.013728
Nature. 2020 Jul;583(7818): 729-736

Global reference mapping of human transcription factor footprints.

Jeff Vierstra, John Lazar, Richard Sandstrom, Jessica Halow, Kristen Lee, Daniel Bates, Morgan Diegel, Douglas Dunn, Fidencio Neri, Eric Haugen, Eric Rynes, Alex Reynolds, Jemma Nelson, Audra Johnson, Mark Frerker, Michael Buckley, Rajinder Kaul, Wouter Meuleman, John A Stamatoyannopoulos.

Combinatorial binding of transcription factors to regulatory DNA underpins gene regulation in all organisms. Genetic variation in regulatory regions has been connected with diseases and diverse phenotypic traits1, but it remains challenging to distinguish variants that affect regulatory function2. Genomic DNase I footprinting enables the quantitative, nucleotide-resolution delineation of sites of transcription factor occupancy within native chromatin3-6. However, only a small fraction of such sites have been precisely resolved on the human genome sequence6. Here, to enable comprehensive mapping of transcription factor footprints, we produced high-density DNase I cleavage maps from 243 human cell and tissue types and states and integrated these data to delineate about 4.5 million compact genomic elements that encode transcription factor occupancy at nucleotide resolution. We map the fine-scale structure within about 1.6 million DNase I-hypersensitive sites and show that the overwhelming majority are populated by well-spaced sites of single transcription factor-DNA interaction. Cell-context-dependent cis-regulation is chiefly executed by wholesale modulation of accessibility at regulatory DNA rather than by differential transcription factor occupancy within accessible elements. We also show that the enrichment of genetic variants associated with diseases or phenotypic traits in regulatory regions1,7 is almost entirely attributable to variants within footprints, and that functional variants that affect transcription factor occupancy are nearly evenly partitioned between loss- and gain-of-function alleles. Unexpectedly, we find increased density of human genetic variation within transcription factor footprints, revealing an unappreciated driver of cis-regulatory evolution. Our results provide a framework for both global and nucleotide-precision analyses of gene regulatory mechanisms and functional genetic variation.

DOI: https://doi.org/10.1038/s41586-020-2528-x
Aging (Albany NY). 2020 Jul 29. 12

HDAC4 inhibition disrupts TET2 function in high-risk MDS and AML.

Feiteng Huang, Jie Sun, Wei Chen, Xin He, Yinghui Zhu, Haojie Dong, Hanying Wang, Zheng Li, Lei Zhang, Samer Khaled, Guido Marcucci, Jinwen Huang, Ling Li.

  Aberrant DNA methylation often silences transcription of tumor-suppressor genes and is considered a hallmark of myeloid neoplasms. Similarly, histone deacetylation represses transcription of genes responsible for cell differentiation/death. A previous clinical study suggested potential pharmacodynamic antagonism between histone deacetylase inhibitors (HDACi) and DNA hypomethylating agents (HMA). Herein, to determine such antagonism, we used MDS/AML lines and NHD13 transgenic mice, and demonstrated that treatment with the pan-HDACi suberoylanilide hydroxamic acid (SAHA) significantly decreased TET2 expression and global 5-hydroxymethylcytosine (5hmC) levels. Mechanistically, our RNAi screen revealed that HDAC4 was responsible for maintaining TET2 levels. Accordingly, HDAC4 knockout reduced expression levels of MTSS1, a known TET2 target, an event associated with decreased 5hmC enrichment on the MTSS1 enhancer. Retrospective analysis of GEO datasets demonstrated that lower HDAC4 levels predict worse prognosis for AML patients. In an MDS-L xenografted immunodeficient mouse model, vitamin C co-treatment prevented TET2 loss of activity seen following SAHA treatment. Accordingly, vitamin C co-treatment further reduced MDS-L cell engraftment relative to SAHA alone. In summary, our findings suggest that co-administration of a TET2 agonist with pan-HDACi treatment could effectively counter potential diminution in TET2 activity resulting from pan-HDACi treatment alone, providing a rationale for evaluating such combinations against high-risk MDS/AML.

Keywords:  TET2; acute myeloid leukemia; histone deacetylase; myelodysplastic syndromes

DOI:  https://doi.org/10.18632/aging.103605
Nucleic Acids Res. 2020 Aug 01. pii: gkaa642. [Epub ahead of print]

Landscape of DNA binding signatures of myocyte enhancer factor-2B reveals a unique interplay of base and shape readout.

Ana Carolina Dantas Machado, Brendon H Cooper, Xiao Lei, Rosa Di Felice, Lin Chen, Remo Rohs.

Myocyte enhancer factor-2B (MEF2B) has the unique capability of binding to its DNA target sites with a degenerate motif, while still functioning as a gene-specific transcriptional regulator. Identifying its DNA targets is crucial given regulatory roles exerted by members of the MEF2 family and MEF2B's involvement in B-cell lymphoma. Analyzing structural data and SELEX-seq experimental results, we deduced the DNA sequence and shape determinants of MEF2B target sites on a high-throughput basis in vitro for wild-type and mutant proteins. Quantitative modeling of MEF2B binding affinities and computational simulations exposed the DNA readout mechanisms of MEF2B. The resulting binding signature of MEF2B revealed distinct intricacies of DNA recognition compared to other transcription factors. MEF2B uses base readout at its half-sites combined with shape readout at the center of its degenerate motif, where A-tract polarity dictates nuances of binding. The predominant role of shape readout at the center of the core motif, with most contacts formed in the minor groove, differs from previously observed protein-DNA readout modes. MEF2B, therefore, represents a unique protein for studies of the role of DNA shape in achieving binding specificity. MEF2B-DNA recognition mechanisms are likely representative for other members of the MEF2 family.

DOI: https://doi.org/10.1093/nar/gkaa642
Nat Commun. 2020 Jul 30. 11(1): 3822

Histone deacetylase 3 controls lung alveolar macrophage development and homeostasis.

Yi Yao, Queping Liu, Indra Adrianto, Xiaojun Wu, James Glassbrook, Namir Khalasawi, Congcong Yin, Qijun Yi, Zheng Dong, Frederic Geissmann, Li Zhou, Qing-Sheng Mi.

Alveolar macrophages (AMs) derived from embryonic precursors seed the lung before birth and self-maintain locally throughout adulthood, but are regenerated by bone marrow (BM) under stress conditions. However, the regulation of AM development and maintenance remains poorly understood. Here, we show that histone deacetylase 3 (HDAC3) is a key epigenetic factor required for AM embryonic development, postnatal homeostasis, maturation, and regeneration from BM. Loss of HDAC3 in early embryonic development affects AM development starting at E14.5, while loss of HDAC3 after birth affects AM homeostasis and maturation. Single-cell RNA sequencing analyses reveal four distinct AM sub-clusters and a dysregulated cluster-specific pathway in the HDAC3-deficient AMs. Moreover, HDAC3-deficient AMs exhibit severe mitochondrial oxidative dysfunction and deteriorative cell death. Mechanistically, HDAC3 directly binds to Pparg enhancers, and HDAC3 deficiency impairs Pparg expression and its signaling pathway. Our findings identify HDAC3 as a key epigenetic regulator of lung AM development and homeostasis.

DOI: https://doi.org/10.1038/s41467-020-17630-6
Elife. 2020 Jul 27. pii: e52483. [Epub ahead of print]9

Dissecting transcriptional amplification by MYC.

Zuqin Nie, Chunhua Guo, Subhendu K Das, Carson C Chow, Eric Batchelor, S Stoney Simons, David Levens.

  Supraphysiological MYC levels are oncogenic. Originally considered a typical transcription factor recruited to E-boxes (CACGTG), another theory posits MYC a global amplifier increasing output at all active promoters. Both models rest on large-scale genome-wide "-omics'. Because the assumptions, statistical parameter and model choice dictates the '-omic' results, whether MYC is a general or specific transcription factor remains controversial. Therefore, an orthogonal series of experiments interrogated MYC's effect on the expression of synthetic reporters. Dose-dependently, MYC increased output at minimal promoters with or without an E-box. Driving minimal promoters with exogenous (glucocorticoid receptor) or synthetic transcription factors made expression more MYC-responsive, effectively increasing MYC-amplifier gain. Mutations of conserved MYC-Box regions I and II impaired amplification, whereas MYC-box III mutations delivered higher reporter output indicating that MBIII limits over-amplification. Kinetic theory and experiments indicate that MYC activates at least two steps in the transcription-cycle to explain the non-linear amplification of transcription that is essential for global, supraphysiological transcription in cancer.

Keywords:  MYC; cancer; cancer biology; computational biology; general amplifier; human; kinetic mechanism of action; michaelis-menten kinetics; systems biology; transcription

DOI:  https://doi.org/10.7554/eLife.52483
Bioinformatics. 2020 Jul 30. pii: btaa687. [Epub ahead of print]

dream: Powerful differential expression analysis for repeated measures designs.

Gabriel E Hoffman, Panos Roussos.

Large-scale transcriptome studies with multiple samples per individual are widely used to study disease biology. Yet current methods for differential expression are inadequate for cross-individual testing for these repeated measures designs. Most problematic, we observe across multiple datasets that current methods can give reproducible false positive findings that are driven by genetic regulation of gene expression, yet are unrelated to the trait of interest. Here we introduce a statistical software package, dream, that increases power, controls the false positive rate, enables multiple types of hypothesis tests, and integrates with standard workflows. In 12 analyses in 6 independent datasets, dream yields biological insight not found with existing software while addressing the issue of reproducible false positive findings.
AVAILABILITY: Dream is available within the variancePartition Bioconductor package at http://bioconductor.org/packages/variancePartition.
SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

DOI: https://doi.org/10.1093/bioinformatics/btaa687