Project description:To understand the mechanism underlying the transcriptional regulation by Sox2, we analyzed genome-wide binding sites of Sox2, Tfap2c, and Cdx2 in trophoblast stem (TS) cells by chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq).
Project description:To understand the mechanism underlying the transcriptional regulation by Sox2, we analyzed genome-wide binding sites of Sox2, Tfap2c, and Cdx2 in trophoblast stem (TS) cells by chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq). ZHBTc4- and embryo-derived TS cell lines were maintained in the presence of FGF4 and mouse embryonic fibroblasts (MEFs).
Project description:Elf5 is a transcription factor with pivotal roles in the trophoblast compartment where it reinforces a trophoblast stem cell (TSC)-specific transcriptional circuit. However, Elf5 is also present in differentiating trophoblast cells that have ceased to express other TSC genes such as Cdx2 and Eomes. In the current study we aimed to elucidate the context-dependent role of Elf5 at the interface between TSC self-renewal and onset of differentiation. We demonstrate that precise levels of Elf5 are critical for normal expansion of the TSC compartment and embryonic survival, as Elf5 overexpression triggers precocious trophoblast differentiation. Through integration of protein interactome, transcriptome and genome-wide chromatin immunoprecipitation data we reveal that this abundance-dependent function is mediated through a shift in preferred Elf5 binding partners; in TSCs, Elf5 interaction with Eomes recruits Tfap2c to triply occupied sites at TSC-specific genes driving their expression. By contrast, the Elf5 and Tfap2c interaction becomes predominant as their protein levels increase. This triggers binding to double and single occupancy sites that harbour the cognate Tfap2c motif, causing activation of the associated differentiation-promoting genes. These data place Elf5 at the centre of a stoichiometry-sensitive transcriptional network where it acts as molecular switch governing the balance between TSC proliferation and differentiation.
Project description:We characterized the trophoblast stem cell epigenome and gene expression profiles in rat and mouse. We profiled 5 histone modifications (+ chromatin input) using ChIP-Seq, and digital expression profiles (3' RNA-Seq) for trophoblast stem cells derived from rat and mouse. Furthermore, for mouse, we profiled key trophoblast stem cell factors Elf5, Cdx2, and Eomes. We found that enhancer regions (defined as distal regions of H3K27ac/H3K4me1 enrichment) were enriched for species-specific endogenous retroviral elements.
Project description:Enhancers are fundamental to gene regulation. Post-translational modifications by the small ubiquitin-like modifiers (SUMO) modify chromatin regulation enzymes, including histone acetylases and deacetylases. However, it remains unclear whether SUMOylation regulates enhancer marks, acetylation at the 27th lysine residue of the histone H3 protein (H3K27Ac). We hypothesize that SUMOylation regulates H3K27Ac. To test this hypothesis, we performed genome-wide ChIP-seq analyses. We discovered that knockdown (KD) of the SUMO activating enzyme catalytic subunit UBA2 reduced H3K27Ac at most enhancers. Bioinformatic analysis revealed that TFAP2C-binding sites are enriched in enhancers whose H3K27Ac was reduced by UBA2 KD. ChIP-seq analysis in combination with molecular biological methods showed that TFAP2C binding to enhancers increased upon UBA2 KD or inhibition of SUMOylation by a small molecule SUMOylation inhibitor. However, this is not due to the SUMOylation of TFAP2C itself. Proteomics analysis of TFAP2C interactome on the chromatin identified histone deacetylation (HDAC) machinery. TFAP2C KD reduced HDAC binding to chromatin and increased H3K27Ac marks at enhancer regions, suggesting that TFAP2C is involved in recruiting HDAC. Taken together, our findings provide important insights into regulation of enhancer marks by SUMOylation.
Project description:The first lineage decisions during mouse development lead to establishment of embryonic and extraembryonic tissues. The transcription factor Cdx2 plays a central role by repressing pluripotency genes, such as Oct4 and promoting trophoblast fate at the blastocyst stage. Here we show that the transcription factor Gata3 is coexpressed with Cdx2 in the blastocyst and that overexpression of Gata3 in embryonic stem cells is sufficient to induce expression of trophoblast genes. Gata3 expression in the blastocyst does not depend on Cdx2, nor do Gata3 overexpressing cell lines require Cdx2 for expression of a subset of trophoblast genes. In the embryo, expression of Gata3, like Cdx2, depends on Tead4, and expression of both factors becomes restricted to nascent trophoblast by an Oct4-independent mechanism. These observations place Tead4 at the top of a trophoblast hierarchy, with Gata3 and Cdx2 acting downstream to induce expression of common and independent targets in this lineage. This SuperSeries is composed of the following subset Series: GSE12985: Differentiation time course of trophoblast stem cells GSE12986: Expression of Cdx2 or Gata3 in R1 mouse embryonic stem cells