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Epigenomic analysis of multilineage differentiation of human embryonic stem cells.

ABSTRACT: 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 nonexpressing 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.

SUBMITTER: Xie W

PROVIDER: S-EPMC3786220 | biostudies-literature | 2013 May

REPOSITORIES: biostudies-literature

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Epigenomic analysis of multilineage differentiation of human embryonic stem cells.

Xie Wei W Schultz Matthew D MD Lister Ryan R Hou Zhonggang Z Rajagopal Nisha N Ray Pradipta P Whitaker John W JW Tian Shulan S Hawkins R David RD Leung Danny D Yang Hongbo H Wang Tao T Lee Ah Young AY Swanson Scott A SA Zhang Jiuchun J Zhu Yun Y Kim Audrey A Nery Joseph R JR Urich Mark A MA Kuan Samantha S Yen Chia-an CA Klugman Sarit S Yu Pengzhi P Suknuntha Kran K Propson Nicholas E NE Chen Huaming H Edsall Lee E LE Wagner Ulrich U Li Yan Y Ye Zhen Z Kulkarni Ashwinikumar A Xuan Zhenyu Z Chung Wen-Yu WY Chi Neil C NC Antosiewicz-Bourget Jessica E JE Slukvin Igor I Stewart Ron R Zhang Michael Q MQ Wang Wei W Thomson James A JA Ecker Joseph R JR Ren Bing B

Cell 20130509 5

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 act ...[more]

PMID: 23664764

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Project description:BackgroundHuman induced pluripotent stem cells (hiPSCs) offer a renewable source of cells for the generation of hematopoietic cells for cell-based therapy, disease modeling, and drug screening. However, current serum/feeder-free differentiation protocols rely on the use of various cytokines, which makes the process very costly or the generation of embryoid bodies (EBs), which are labor-intensive and can cause heterogeneity during differentiation. Here, we report a simple feeder and serum-free monolayer protocol for efficient generation of iPSC-derived multipotent hematoendothelial progenitors (HEPs), which can further differentiate into endothelial and hematopoietic cells including erythroid and T lineages.MethodsFormation of HEPs from iPSCs was initiated by inhibition of GSK3 signaling for 2 days followed by the addition of VEGF and FGF2 for 3 days. The HEPs were further induced toward mature endothelial cells (ECs) in an angiogenic condition and toward T cells by co-culturing with OP9-DL1 feeder cells. Endothelial-to-hematopoietic transition (EHT) of the HEPs was further promoted by supplementation with the TGF-β signaling inhibitor. Erythroid differentiation was performed by culturing the hematopoietic stem/progenitor cells (HSPCs) in a three-stage erythroid liquid culture system.ResultsOur protocol significantly enhanced the number of KDR+ CD34+ CD31+ HEPs on day 5 of differentiation. Further culture of HEPs in angiogenic conditions promoted the formation of mature ECs, which expressed CD34, CD31, CD144, vWF, and ICAM-1, and could exhibit the formation of vascular-like network and acetylated low-density lipoprotein (Ac-LDL) uptake. In addition, the HEPs were differentiated into CD8+ T lymphocytes, which could be expanded up to 34-fold upon TCR stimulation. Inhibition of TGF-β signaling at the HEP stage promoted EHT and yielded a large number of HSPCs expressing CD34 and CD43. Upon erythroid differentiation, these HSPCs were expanded up to 40-fold and displayed morphological changes following stages of erythroid development.ConclusionThis protocol offers an efficient and simple approach for the generation of multipotent HEPs and could be adapted to generate desired blood cells in large numbers for applications in basic research including developmental study, disease modeling, and drug screening as well as in regenerative medicine.

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Epigenomic analysis of multilineage differentiation of human embryonic stem cells.

Publications

Epigenomic analysis of multilineage differentiation of human embryonic stem cells.

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