Transcriptional and epigenetic dynamics observed during lineage specification of human embryonic stem cells.

摘要

Epigenetic regulation of gene expression is essential for faithful cellular specification during embryonic development. Directed differentiation of pluripotent human embryonic stem cells (hESCs), which maintain the ability to give rise to each cell type found within the human body, provides a tractable system to study both the epigenetic mechanisms that facilitate cellular transitions, and the transcription factors (TFs) that dictate these events. To understand molecular events associated with major lineage decisions, we performed comprehensive genomic profiling, including RNA-Sequencing, Chromatin Immunoprecipitation-Sequencing (ChIP-Seq) for six histone modifications and whole genome bisulfite-sequencing (WGBS) to interrogate DNA methylation levels, on three populations derived through directed differentiation of hESCs. Expression profiling detected signatures that resembled the three embryonic germ layers, namely ectoderm, mesoderm and endoderm. Integration of ChIP-Seq and WGBS data revealed widespread remodeling, predominantly at intergenic regions. To understand the impact of TF binding on epigenetic remodeling, we then complemented the epigenetic information with binding profiles for the pluripotency TFs OCT4, SOX2 and NANOG (O/S/N) in hESCs, and FOXA2 in the endoderm population. O/S/N binding was identified near pluripotency genes as expected, as well as regions that exhibited lineage specific remodeling during differentiation and are linked to later stages of development. We also discerned a novel epigenetic trend, in which H3K27me3 was unexpectedly gained at regions of low CpG density that exhibit high levels of DNA methylation in hESCs. These events overlapped with FOXA2 binding sites in the dEN that lose DNA methylation. Notably, these events were detected near genes associated with later stages of development, such as AFP. We postulate that these FOXA2-associated epigenetic remodeling events lead to acquisition of a transient, facultative heterochromatic state necessary to foster efficient differentiation of subsequent stages. Integration of these data sets yielded an unprecedented perspective of the orchestrated transcriptional and epigenetic events that occur during cell state transitions. Future studies that compare epigenomic profiles of in vitro derived cell types to their primary counterparts may identify regulatory elements that are held in improper epigenetic states, and ultimately lead to improved differentiation protocols and the in vitro derivation of therapeutically relevant cell types.