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Current catalogs of regulatory sequences in the human genome are still incomplete and lack cell type resolution. To profile the activity of gene regulatory elements in diverse cell types and tissues in the human body, we applied single-cell chromatin accessibility assays to 30 adult human tissue types from multiple donors. We integrated these datasets with previous single-cell chromatin accessibility data from 15 fetal tissue types to reveal the status of open chromatin for ∼1.2 million candidate cis-regulatory elements (cCREs) in 222 distinct cell types comprised of >1.3 million nuclei. We used these chromatin accessibility maps to delineate cell-type-specificity of fetal and adult human cCREs and to systematically interpret the noncoding variants associated with complex human traits and diseases. This rich resource provides a foundation for the analysis of gene regulatory programs in human cell types across tissues, life stages, and organ systems., Competing Interests: Declaration of interests B.R. is a shareholder and consultant of Arima Genomics, Inc., and a co-founder of Epigenome Technologies, Inc. K.J.G. is a consultant of Genentech and a shareholder in Vertex Pharmaceuticals. These relationships have been disclosed to and approved by the UCSD Independent Review Committee., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Single-nucleus assay for transposase-accessible chromatin using sequencing (snATAC-seq) creates new opportunities to dissect cell type-specific mechanisms of complex diseases. Since pancreatic islets are central to type 2 diabetes (T2D), we profiled 15,298 islet cells by using combinatorial barcoding snATAC-seq and identified 12 clusters, including multiple alpha, beta and delta cell states. We cataloged 228,873 accessible chromatin sites and identified transcription factors underlying lineage- and state-specific regulation. We observed state-specific enrichment of fasting glucose and T2D genome-wide association studies for beta cells and enrichment for other endocrine cell types. At T2D signals localized to islet-accessible chromatin, we prioritized variants with predicted regulatory function and co-accessibility with target genes. A causal T2D variant rs231361 at the KCNQ1 locus had predicted effects on a beta cell enhancer co-accessible with INS and genome editing in embryonic stem cell-derived beta cells affected INS levels. Together our findings demonstrate the power of single-cell epigenomics for interpreting complex disease genetics.

Background: Co-localized combinations of histone modifications ("chromatin states") have been shown to correlate with promoter and enhancer activity. Changes in chromatin states over multiple time points ("chromatin state trajectories") have previously been analyzed at promoter and enhancers separately. With the advent of time series Hi-C data it is now possible to connect promoters and enhancers and to analyze chromatin state trajectories at promoter-enhancer pairs., Results: We present TimelessFlex, a framework for investigating chromatin state trajectories at promoters and enhancers and at promoter-enhancer pairs based on Hi-C information. TimelessFlex extends our previous approach Timeless, a Bayesian network for clustering multiple histone modification data sets at promoter and enhancer feature regions. We utilize time series ATAC-seq data measuring open chromatin to define promoters and enhancer candidates. We developed an expectation-maximization algorithm to assign promoters and enhancers to each other based on Hi-C interactions and jointly cluster their feature regions into paired chromatin state trajectories. We find jointly clustered promoter-enhancer pairs showing the same activation patterns on both sides but with a stronger trend at the enhancer side. While the promoter side remains accessible across the time series, the enhancer side becomes dynamically more open towards the gene activation time point. Promoter cluster patterns show strong correlations with gene expression signals, whereas Hi-C signals get only slightly stronger towards activation. The code of the framework is available at https://github.com/henriettemiko/TimelessFlex ., Conclusions: TimelessFlex clusters time series histone modifications at promoter-enhancer pairs based on Hi-C and it can identify distinct chromatin states at promoter and enhancer feature regions and their changes over time.

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.

Chromatin architecture has been implicated in cell type-specific gene regulatory programs, yet how chromatin remodels during development remains to be fully elucidated. Here, by interrogating chromatin reorganization during human pluripotent stem cell (hPSC) differentiation, we discover a role for the primate-specific endogenous retrotransposon human endogenous retrovirus subfamily H (HERV-H) in creating topologically associating domains (TADs) in hPSCs. Deleting these HERV-H elements eliminates their corresponding TAD boundaries and reduces the transcription of upstream genes, while de novo insertion of HERV-H elements can introduce new TAD boundaries. The ability of HERV-H to create TAD boundaries depends on high transcription, as transcriptional repression of HERV-H elements prevents the formation of boundaries. This ability is not limited to hPSCs, as these actively transcribed HERV-H elements and their corresponding TAD boundaries also appear in pluripotent stem cells from other hominids but not in more distantly related species lacking HERV-H elements. Overall, our results provide direct evidence for retrotransposons in actively shaping cell type- and species-specific chromatin architecture.

Genetic variants affecting pancreatic islet enhancers are central to T2D risk, but the gene targets of islet enhancer activity are largely unknown. We generate a high-resolution map of islet chromatin loops using Hi-C assays in three islet samples and use loops to annotate target genes of islet enhancers defined using ATAC-seq and published ChIP-seq data. We identify candidate target genes for thousands of islet enhancers, and find that enhancer looping is correlated with islet-specific gene expression. We fine-map T2D risk variants affecting islet enhancers, and find that candidate target genes of these variants defined using chromatin looping and eQTL mapping are enriched in protein transport and secretion pathways. At IGF2BP2, a fine-mapped T2D variant reduces islet enhancer activity and IGF2BP2 expression, and conditional inactivation of IGF2BP2 in mouse islets impairs glucose-stimulated insulin secretion. Our findings provide a resource for studying islet enhancer function and identifying genes involved in T2D risk.

Long-range chromatin interactions between enhancers and promoters are essential for transcription of many developmentally controlled genes in mammals and other metazoans. Currently, the exact mechanisms that connect distal enhancers to their specific target promoters remain to be fully elucidated. Here, we show that the enhancer-specific histone H3 lysine 4 monomethylation (H3K4me1) and the histone methyltransferases MLL3 and MLL4 (MLL3/4) play an active role in this process. We demonstrate that in differentiating mouse embryonic stem cells, MLL3/4-dependent deposition of H3K4me1 at enhancers correlates with increased levels of chromatin interactions, whereas loss of this histone modification leads to reduced levels of chromatin interactions and defects in gene activation during differentiation. H3K4me1 facilitates recruitment of the Cohesin complex, a known regulator of chromatin organization, to chromatin in vitro and in vivo, providing a potential mechanism for MLL3/4 to promote chromatin interactions between enhancers and promoters. Taken together, our results support a role for MLL3/4-dependent H3K4me1 in orchestrating long-range chromatin interactions at enhancers in mammalian cells.

The three-dimensional configuration of DNA is integral to all nuclear processes in eukaryotes, yet our knowledge of the chromosome architecture is still limited. Genome-wide chromosome conformation capture studies have uncovered features of chromatin organization in cultured cells, but genome architecture in human tissues has yet to be explored. Here, we report the most comprehensive survey to date of chromatin organization in human tissues. Through integrative analysis of chromatin contact maps in 21 primary human tissues and cell types, we find topologically associating domains highly conserved in different tissues. We also discover genomic regions that exhibit unusually high levels of local chromatin interactions. These frequently interacting regions (FIREs) are enriched for super-enhancers and are near tissue-specifically expressed genes. They display strong tissue-specificity in local chromatin interactions. Additionally, FIRE formation is partially dependent on CTCF and the Cohesin complex. We further show that FIREs can help annotate the function of non-coding sequence variants., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

The mammalian cerebrum performs high-level sensory perception, motor control and cognitive functions through highly specialized cortical and subcortical structures1. Recent surveys of mouse and human brains with single-cell transcriptomics2-6 and high-throughput imaging technologies7,8 have uncovered hundreds of neural cell types distributed in different brain regions, but the transcriptional regulatory programs that are responsible for the unique identity and function of each cell type remain unknown. Here we probe the accessible chromatin in more than 800,000 individual nuclei from 45 regions that span the adult mouse isocortex, olfactory bulb, hippocampus and cerebral nuclei, and use the resulting data to map the state of 491,818 candidate cis-regulatory DNA elements in 160 distinct cell types. We find high specificity of spatial distribution for not only excitatory neurons, but also most classes of inhibitory neurons and a subset of glial cell types. We characterize the gene regulatory sequences associated with the regional specificity within these cell types. We further link a considerable fraction of the cis-regulatory elements to putative target genes expressed in diverse cerebral cell types and predict transcriptional regulators that are involved in a broad spectrum of molecular and cellular pathways in different neuronal and glial cell populations. Our results provide a foundation for comprehensive analysis of gene regulatory programs of the mammalian brain and assist in the interpretation of noncoding risk variants associated with various neurological diseases and traits in humans.

Genetic variants associated with type 2 diabetes (T2D) risk affect gene regulation in metabolically relevant tissues, such as pancreatic islets. Here, we investigated contributions of regulatory programs active during pancreatic development to T2D risk. Generation of chromatin maps from developmental precursors throughout pancreatic differentiation of human embryonic stem cells (hESCs) identifies enrichment of T2D variants in pancreatic progenitor-specific stretch enhancers that are not active in islets. Genes associated with progenitor-specific stretch enhancers are predicted to regulate developmental processes, most notably tissue morphogenesis. Through gene editing in hESCs, we demonstrate that progenitor-specific enhancers harboring T2D-associated variants regulate cell polarity genes LAMA1 and CRB2. Knockdown of lama1 or crb2 in zebrafish embryos causes a defect in pancreas morphogenesis and impairs islet cell development. Together, our findings reveal that a subset of T2D risk variants specifically affects pancreatic developmental programs, suggesting that dysregulation of developmental processes can predispose to T2D.

In meiotic prophase, chromosomes are organized into compacted loop arrays to promote homolog pairing and recombination. Here, we probe the architecture of the mouse spermatocyte genome in early and late meiotic prophase using chromosome conformation capture (Hi-C). Our data support the established loop array model of meiotic chromosomes, and infer loops averaging 0.8-1.0 megabase pairs (Mb) in early prophase and extending to 1.5-2.0 Mb in late prophase as chromosomes compact and homologs undergo synapsis. Topologically associating domains (TADs) are lost in meiotic prophase, suggesting that assembly of the meiotic chromosome axis alters the activity of chromosome-associated cohesin complexes. While TADs are lost, physically separated A and B compartments are maintained in meiotic prophase. Moreover, meiotic DNA breaks and interhomolog crossovers preferentially form in the gene-dense A compartment, revealing a role for chromatin organization in meiotic recombination. Finally, direct detection of interhomolog contacts genome-wide reveals the structural basis for homolog alignment and juxtaposition by the synaptonemal complex.

FOXA pioneer transcription factors (TFs) associate with primed enhancers in endodermal organ precursors. Using a human stem cell model of pancreas differentiation, we here discover that only a subset of pancreatic enhancers is FOXA-primed, whereas the majority is unprimed and engages FOXA upon lineage induction. Primed enhancers are enriched for signal-dependent TF motifs and harbor abundant and strong FOXA motifs. Unprimed enhancers harbor fewer, more degenerate FOXA motifs, and FOXA recruitment to unprimed but not primed enhancers requires pancreatic TFs. Strengthening FOXA motifs at an unprimed enhancer near NKX6.1 renders FOXA recruitment pancreatic TF-independent, induces priming, and broadens the NKX6.1 expression domain. We make analogous observations about FOXA binding during hepatic and lung development. Our findings suggest a dual role for FOXA in endodermal organ development: first, FOXA facilitates signal-dependent lineage initiation via enhancer priming, and second, FOXA enforces organ cell type-specific gene expression via indirect recruitment by lineage-specific TFs. Enhancers for endodermal organs are primed at the chromatin level prior to lineage induction by FOXA pioneer transcription factors; how pervasive this is, is not well known. Here the authors show that only a small subset of organ-specific enhancers are bound and primed by FOXA prior to lineage induction, whereas the majority do not undergo chromatin priming and engage FOXA upon lineage induction. [ABSTRACT FROM AUTHOR]

Hi-C and chromatin immunoprecipitation (ChIP) have been combined to identify long-range chromatin interactions genome-wide at reduced cost and enhanced resolution, but extracting information from the resulting datasets has been challenging. Here we describe a computational method, MAPS, Model-based Analysis of PLAC-seq and HiChIP, to process the data from such experiments and identify long-range chromatin interactions. MAPS adopts a zero-truncated Poisson regression framework to explicitly remove systematic biases in the PLAC-seq and HiChIP datasets, and then uses the normalized chromatin contact frequencies to identify significant chromatin interactions anchored at genomic regions bound by the protein of interest. MAPS shows superior performance over existing software tools in the analysis of chromatin interactions from multiple PLAC-seq and HiChIP datasets centered on different transcriptional factors and histone marks. MAPS is freely available at . [ABSTRACT FROM AUTHOR]