Unknown,Transcriptomics,Genomics,Proteomics

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Transcription factor binding dynamics during human ES cell differentiation

ABSTRACT: Pluripotent stem cells provide a powerful system to dissect the underlying molecular dynamics that regulate cell fate changes during mammalian development. Here we report the integrative analysis of genome wide binding data for 38 transcription factors with extensive epigenome and transcriptional data across the differentiation of human embryonic stem cells to the three germ layers. We describe core regulatory dynamics and show the lineage specific behavior of selected factors. In addition to the orchestrated remodeling of the chromatin landscape, we find that the binding of several transcription factors is strongly associated with specific loss of DNA methylation in one germ layer and in many cases a reciprocal gain in the other layers. Taken together, our work shows context-dependent rewiring of transcription factor binding, downstream signaling effectors, and the epigenome during human embryonic stem cell differentiation. 200 ChIP-seq experiments profiling 38 transcription factors (TFs) and several chromatin marks in 5 cell types--male human ES cell line HUES64 and directed differentiation of HUES64 towards mesendoderm (dMS, 12 hours), endoderm (dEN, 120 hours), mesoderm (dME, 120 hours), and ectoderm (dEC, 120 hours). In addition, three ES cell lines were derived with shRNA mediated knockdown of GATA4 and differention toward endoderm (dEN_shGATA4) and mesoderm (dME_shGATA4). These cell lines were used for MNChIP-seq of GATA4, SMAD1, and H3K27Ac and for 4 RRBS experiments in GATA4 knockdown and control cell lines.

ORGANISM(S): Homo sapiens

SUBMITTER: Alexander Tsankov

PROVIDER: E-GEOD-61475 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Transcription factor binding dynamics during human ES cell differentiation.

Tsankov Alexander M AM Gu Hongcang H Akopian Veronika V Ziller Michael J MJ Donaghey Julie J Amit Ido I Gnirke Andreas A Meissner Alexander A

Nature 20150201 7539

Pluripotent stem cells provide a powerful system to dissect the underlying molecular dynamics that regulate cell fate changes during mammalian development. Here we report the integrative analysis of genome-wide binding data for 38 transcription factors with extensive epigenome and transcriptional data across the differentiation of human embryonic stem cells to the three germ layers. We describe core regulatory dynamics and show the lineage-specific behaviour of selected factors. In addition to t ...[more]

PMID: 25693565

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Project description:We show that high quality microarray gene expression profiles can be obtained following FACS sorting of cells using combinations of transcription factors. We use this transcription factor FACS (tfFACS) methodology to perform a genomic analysis of hESC-derived endodermal lineages marked by combinations of SOX17, GATA4, and CXCR4, and find that triple positive cells have a much stronger definitive endoderm signature than other combinations of these markers. Additionally, SOX17+GATA4+ cells can be obtained at a much earlier stage of differentiation, prior to expression of CXCR4+ cells, providing an important new tool to isolate this earlier definitive endoderm subtype. Overall, tfFACS represents an advancement in FACS technology which broadly crosses multiple disciplines, most notably in regenerative medicine to redefine cellular populations. 21 Total samples were analyzed. Samples collected after 5 days of hESC differentiation included SOX17+GATA4+ CXCR4+ cells, unfixed CXCR4+ cells, and unsorted fixed cells. Samples collected after 3 days of differentiation included SOX17+GATA4+ cells, SOX17-GATA- cells, and unsorted fixed cells. As controls, we analyzed both unfixed hESCs and fixed hESCs. All samples contained biological duplicates, triplicates or quadriplicates. We performed hierarchical clustering to demonstrate whether cellular fixation alone could change gene expression. We based this analysis on 1647 transcript clusters with coefficient of variation > 0.5 across the samples and expression values >= 500 in at least 2 out of the 21 samples. We found that fixation and staining steps did not cause distortions in gene expression measurements. This is supported by the fact that fixed and unfixed cells cluster together based upon differentiation stage, not based upon degree of fixation. Furthermore, using GSEA analysis, we found that the SOX17+GATA4+CXCR4+ day 5 cells and day 3 SOX17+GATA4+ were more enriched for compiled gold-standard endodermal genes than the CXCR4+ day 5 population.

Dataset Information

Transcription factor binding dynamics during human ES cell differentiation

Publications

Transcription factor binding dynamics during human ES cell differentiation.

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