Integration of External Signaling Pathways with the Core Transcriptional Network in Embryonic Stem Cells

Abstract

Transcription factors (TFs) and their specific interactions with targets are crucial for specifying gene-expression programs. To gain insights into the transcriptional regulatory networks in embryonic stem (ES) cells, we use chromatin immunoprecipitation coupled with ultra-high-throughput DNA sequencing (ChIP-seq) to map the locations of 13 sequence-specific TFs (Nanog, Oct4, STAT3, Smad1, Sox2, Zfx, c-Myc, n-Myc, Klf4, Esrrb, Tcfcp2l1, E2f1, and CTCF) and 2 transcription regulators (p300 and Suz12). These factors are known to play different roles in ES-cell biology as components of the LIF and BMP signaling pathways, self-renewal regulators, and key reprogramming factors. Our study provides insights into the integration of the signaling pathways into the ES-cell-specific transcription circuitries. Intriguingly, we find specific genomic regions extensively targeted by different TFs. Collectively, the comprehensive mapping of TF-binding sites identifies important features of the transcriptional regulatory networks that define ES-cell identity.

Esrrb extinction triggers dismantling of naïve pluripotency and marks commitment to differentiation

“…Comparison with publicly available TF‐binding datasets in ESCs (Chen et al , 2008) revealed that Class I elements are bound by ESRRB, KLF4, and NANOG more robustly than Class II elements (Fig 6A). In contrast, Class I elements bind OCT4 less than Class II elements (Fig 6A).…”

“…Data for all peaks near genes measured in our microarray assay are shown for comparison [gray dashed line; data from Chen et al (2008) and Kagey et al (2010)]. Read counts per base pair were normalized by library size and by subtracting the background signal (IgG).…”

“…Microarray analyses further indicated that the majority of known NANOG and ESRRB target genes (Festuccia et al , 2012) are among the genes with the most dramatic change in expression during the transition from Esrrb‐GFP high/medium to Esrrb‐GFP negative cells (55.6% of NANOG or ESRRB targets are in the top/bottom 400 genes ranked by the expression difference between Esrrb‐GFP high and Esrrb‐GFP negative cells; hypergeometric P  = 1.22 × 10 −35 ; Fig 3F, Table EV1). Analysis of the regulatory regions in proximity of Differentially Expressed Genes (DEGs) in publicly available ChIP‐Seq datasets showed enrichment for binding of NANOG, ESRRB, KLF4, and the mediator subunit MED1 (Chen et al , 2008; Fig EV3B), suggesting that these DEGs are stringently controlled by enhancers through which naïve pluripotency TFs act. Taken together, these results suggest that modulation of the entire transcriptional programs controlled by NANOG and ESRRB accompanies commitment to differentiation.…”

“…Error bars: standard deviation of the measures in four independent experiments. ChIP‐Seq read coverage for the indicated factors derived from re‐analysis of published datasets (Chen et al , 2008) shows how TF binding correlates with methylation “valleys” and increased chromatin accessibility. RNA polymerase II binding marks promoters (Stadler et al , 2011).…”

Modeling heterogeneity in the pluripotent state: A promising strategy for improving the efficiency and fidelity of stem cell differentiation

“…The TGF‐β/SMAD signaling pathway maintains self‐renewal in mESC through the BMP/SMAD signalling activation of Id family genes 52, while in hESCs is the Activin/Nodal/SMAD2/3 cascade the one responsible for promoting pluripotency 53. Interestingly, several studies have demonstrated that pluripotency regulators form intricate circuits at the transcriptional level 13, 36, 53, 54, and many of the TFs ( Essrb , Klf4 , Stat3 , Tcf3 ) in these regulatory motifs are downstream effectors of the signaling pathways regulating self‐renewal and differentiation. Although the complete spectrum of signaling pathways regulating pluripotency has not been fully described 55, these results demonstrate the confluence of different environmental signals from the microenvironment for the regulation of pluripotency.…”