The reference human genome sequence set the stage for studies of genetic variation and its association with human disease, but a similar reference has lacked for epigenomic studies. To address this need, the NIH Roadmap Epigenomics Consortium generated the largest collection to-date of human epigenomes for primary cells and tissues. Here, we describe the integrative analysis of 111 reference human epigenomes generated as part of the program, profiled for histone modification patterns, DNA accessibility, DNA methylation, and RNA expression. We establish global maps of regulatory elements, define regulatory modules of coordinated activity, and their likely activators and repressors. We show that disease and trait-associated genetic variants are enriched in tissue-specific epigenomic marks, revealing biologically-relevant cell types for diverse human traits, and providing a resource for interpreting the molecular basis of human disease. Our results demonstrate the central role of epigenomic information for understanding gene regulation, cellular differentiation, and human disease.
Higher order chromatin structure is emerging as an important regulator of gene expression. Although dynamic chromatin structures have been identified in the genome, the full scope of chromatin dynamics during mammalian development and lineage specification remains obscure. By mapping genome-wide chromatin interactions in human embryonic stem cells (hESC) and four hESC-derived lineages, we uncover extensive chromatin reorganization during lineage specification. We observe that while topological domain boundaries remain intact during differentiation, interactions both within and between domains change dramatically, altering 36% of active and inactive chromosomal “compartments” throughout the genome. By integrating chromatin interaction maps with haplotype-resolved epigenome and transcriptome datasets, we find widespread allelic bias in gene expression correlated with allele-biased chromatin states of linked promoters and distal enhancers. Our results therefore provide a global view of chromatin dynamics and a resource for studying long-range control of gene expression in distinct human cell lineages.
Ectopic expression of the four transcription factors Oct4, Sox2, c-Myc, and Klf4 is sufficient to confer a pluripotent state upon the fibroblast genome, generating induced pluripotent stem (iPS) cells. It remains unknown if nuclear reprogramming induced by these four factors globally resets epigenetic differences between differentiated and pluripotent cells. Here, using novel selection approaches, we have generated iPS cells from fibroblasts to characterize their epigenetic state. Female iPS cells showed reactivation of a somatically silenced X chromosome and underwent random X inactivation upon differentiation. Genome-wide analysis of two key histone modifications indicated that iPS cells are highly similar to ES cells. Consistent with these observations, iPS cells gave rise to viable high-degree chimeras with contribution to the germline. These data show that transcription factor-induced reprogramming leads to the global reversion of the somatic epigenome into an ES-like state. Our results provide a paradigm for studying the epigenetic modifications that accompany nuclear reprogramming and suggest that abnormal epigenetic reprogramming does not pose a problem for the potential therapeutic applications of iPS cells.
SUMMARY Induced pluripotent stem cells (iPSCs) outwardly appear to be indistinguishable from embryonic stem cells (ESCs). A study of gene expression profiles of mouse and human ESCs and iPSCs suggests that, while iPSCs are quite similar to their embryonic counterparts, a recurrent gene expression signature appears in iPSCs regardless of their origin or the method by which they were generated. Upon extended culture, hiPSCs adopt a gene expression profile more similar to hESCs; however, they still retain a gene expression signature unique from hESCs that extends to miRNA expression. Genome-wide data suggested that the iPSC signature gene expression differences are due to differential promoter binding by the reprogramming factors. High-resolution array profiling demonstrated that there is no common specific subkaryotypic alteration that is required for reprogramming and that reprogramming does not lead to genomic instability. Together, these data suggest that iPSCs should be considered a unique subtype of pluripotent cell.
SUMMARY Epigenetic mechanisms have been proposed to play crucial roles in mammalian development, but their precise functions are only partially understood. To investigate epigenetic regulation of embryonic development, we differentiated human embryonic stem cells into mesendoderm, neural progenitor cells, trophoblast-like cells, and mesenchymal stem cells, and systematically characterized DNA methylation, chromatin modifications, and the transcriptome in each lineage. We found that promoters that are active in early developmental stages tend to be CG rich and mainly engage H3K27me3 upon silencing in non-expressing lineages. By contrast, promoters for genes expressed preferentially at later stages are often CG poor and primarily employ DNA methylation upon repression. Interestingly, the early developmental regulatory genes are often located in large genomic domains that are generally devoid of DNA methylation in most lineages, which we termed DNA methylation valleys (DMVs). Our results suggest that distinct epigenetic mechanisms regulate early and late stages of ES cell differentiation.
Histone modifications are fundamental epigenetic regulators that control many crucial cellular processes. However, whether these marks can be passed on from mammalian gametes to the next generation is a long-standing question that remains unanswered. Here, by developing a highly sensitive approach, STAR ChIP-seq, we provide a panoramic view of the landscape of H3K4me3, a histone hallmark for transcription initiation, from developing gametes to post-implantation embryos. We find that upon fertilization, extensive reprogramming occurs on the paternal genome, as H3K4me3 peaks are depleted in zygotes but are readily observed after major zygotic genome activation at the late two-cell stage. On the maternal genome, we unexpectedly find a non-canonical form of H3K4me3 (ncH3K4me3) in full-grown and mature oocytes, which exists as broad peaks at promoters and a large number of distal loci. Such broad H3K4me3 peaks are in contrast to the typical sharp H3K4me3 peaks restricted to CpG-rich regions of promoters. Notably, ncH3K4me3 in oocytes overlaps almost exclusively with partially methylated DNA domains. It is then inherited in pre-implantation embryos, before being erased in the late two-cell embryos, when canonical H3K4me3 starts to be established. The removal of ncH3K4me3 requires zygotic transcription but is independent of DNA replication-mediated passive dilution. Finally, downregulation of H3K4me3 in full-grown oocytes by overexpression of the H3K4me3 demethylase KDM5B is associated with defects in genome silencing. Taken together, these data unveil inheritance and highly dynamic reprogramming of the epigenome in early mammalian development.
In mammals, extensive chromatin reorganization is essential for reprogramming terminally committed gametes to a totipotent state during preimplantation development. However, the global chromatin landscape and its dynamics in this period remain unexplored. Here we report a genome-wide map of accessible chromatin in mouse preimplantation embryos using an improved assay for transposase-accessible chromatin with high throughput sequencing (ATAC-seq) approach with CRISPR/Cas9-assisted mitochondrial DNA depletion. We show that despite extensive parental asymmetry in DNA methylomes, the chromatin accessibility between the parental genomes is globally comparable after major zygotic genome activation (ZGA). Accessible chromatin in early embryos is widely shaped by transposable elements and overlaps extensively with putative cis-regulatory sequences. Unexpectedly, accessible chromatin is also found near the transcription end sites of active genes. By integrating the maps of cis-regulatory elements and single-cell transcriptomes, we construct the regulatory network of early development, which helps to identify the key modulators for lineage specification. Finally, we find that the activities of cis-regulatory elements and their associated open chromatin diminished before major ZGA. Surprisingly, we observed many loci showing non-canonical, large open chromatin domains over the entire transcribed units in minor ZGA, supporting the presence of an unusually permissive chromatin state. Together, these data reveal a unique spatiotemporal chromatin configuration that accompanies early mammalian development.
In mammals, chromatin organization undergoes drastic reprogramming after fertilization. However, the three-dimensional structure of chromatin and its reprogramming in preimplantation development remain poorly understood. Here, by developing a low-input Hi-C (genome-wide chromosome conformation capture) approach, we examined the reprogramming of chromatin organization during early development in mice. We found that oocytes in metaphase II show homogeneous chromatin folding that lacks detectable topologically associating domains (TADs) and chromatin compartments. Strikingly, chromatin shows greatly diminished higher-order structure after fertilization. Unexpectedly, the subsequent establishment of chromatin organization is a prolonged process that extends through preimplantation development, as characterized by slow consolidation of TADs and segregation of chromatin compartments. The two sets of parental chromosomes are spatially separated from each other and display distinct compartmentalization in zygotes. Such allele separation and allelic compartmentalization can be found as late as the 8-cell stage. Finally, we show that chromatin compaction in preimplantation embryos can partially proceed in the absence of zygotic transcription and is a multi-level hierarchical process. Taken together, our data suggest that chromatin may exist in a markedly relaxed state after fertilization, followed by progressive maturation of higher-order chromatin architecture during early development.
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