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Identification of transcription factors that interact with Arid4b protein in mouse embryonic stem cells

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2019
Keskin, Ezgi Gül
Embryonic development is a process that is highly regulated at the level of gene expression. Throughout this process, the zygote divides and differentiates to form all cell types in the adult body. In the early stages of development, pluripotent embryonic stem cells (ESC) can be isolated and used to study cell fate mechanisms during development in vitro. The pluripotency of ESCs is governed by certain transcription factors (TF) (Oct4, Nanog, etc.). These TFs construct a transcription network specific to ESC and enable the high and stable expression of pluripotency genes. Chromatin factors work alongside TFs to maintain pluripotency. In our previous studies, we have identified Arid4b protein in the Sin3a corepressor complex is critical for proper ESC differentiation into mesoderm and endoderm lineages. Arid4b might direct the Sin3a complex to the ESC differentiation related genes with the aid of its histone modification and DNA binding domains. Histone deacetylases in the complex is critical for the suppression of gene expression. To better understand the role of Arid4b protein in ESC differentiation, we examined the histone 3 lysine 27 acetylation (H3K27Ac) genome-wide distribution in wildtype and arid4bΔ mesoderm or endoderm differentiated cells. H3K27Ac is a marker of active enhancer regions as well as active promotor regions. We found that the higher level of H3K27Ac in the arid4bΔ cells when compared to wildtype cells. Therefore, Sin3a complex may be required for the suppression of these genes during ESC differentiation. Upon bioinformatics analyses, we identified TF consensus binding sequences enriched in the regions with increased H3K27Ac levels in arid4bΔ cells. We analyzed the mRNA expression level of 10 such TFs in WT and arid4bΔ ESCs. We further validated the protein expression of selected TFs. Of these, we showed that Tfap2c physically interacts with Arid4b and Sin3a in ESCs through endogenous coimmunoprecipitation experiments.