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Brg1 is required for Cdx2-mediated repression of Oct4 expression in mouse blastocysts.

ABSTRACT: During blastocyst formation the segregation of the inner cell mass (ICM) and trophectoderm is governed by the mutually antagonistic effects of the transcription factors Oct4 and Cdx2. Evidence indicates that suppression of Oct4 expression in the trophectoderm is mediated by Cdx2. Nonetheless, the underlying epigenetic modifiers required for Cdx2-dependent repression of Oct4 are largely unknown. Here we show that the chromatin remodeling protein Brg1 is required for Cdx2-mediated repression of Oct4 expression in mouse blastocysts. By employing a combination of RNA interference (RNAi) and gene expression analysis we found that both Brg1 Knockdown (KD) and Cdx2 KD blastocysts exhibit widespread expression of Oct4 in the trophectoderm. Interestingly, in Brg1 KD blastocysts and Cdx2 KD blastocysts, the expression of Cdx2 and Brg1 is unchanged, respectively. To address whether Brg1 cooperates with Cdx2 to repress Oct4 transcription in the developing trophectoderm, we utilized preimplantation embryos, trophoblast stem (TS) cells and Cdx2-inducible embryonic stem (ES) cells as model systems. We found that: (1) combined knockdown (KD) of Brg1 and Cdx2 levels in blastocysts resulted in increased levels of Oct4 transcripts compared to KD of Brg1 or Cdx2 alone, (2) endogenous Brg1 co-immunoprecipitated with Cdx2 in TS cell extracts, (3) in blastocysts Brg1 and Cdx2 co-localize in trophectoderm nuclei and (4) in Cdx2-induced ES cells Brg1 and Cdx2 are recruited to the Oct4 promoter. Lastly, to determine how Brg1 may induce epigenetic silencing of the Oct4 gene, we evaluated CpG methylation at the Oct4 promoter in the trophectoderm of Brg1 KD blastocysts. This analysis revealed that Brg1-dependent repression of Oct4 expression is independent of DNA methylation at the blastocyst stage. In toto, these results demonstrate that Brg1 cooperates with Cdx2 to repress Oct4 expression in the developing trophectoderm to ensure normal development.

SUBMITTER: Wang K

PROVIDER: S-EPMC2868905 | biostudies-literature | 2010 May

REPOSITORIES: biostudies-literature

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Brg1 is required for Cdx2-mediated repression of Oct4 expression in mouse blastocysts.

Wang Kai K Sengupta Satyaki S Magnani Luca L Wilson Catherine A CA Henry R William RW Knott Jason G JG

PloS one 20100512 5

During blastocyst formation the segregation of the inner cell mass (ICM) and trophectoderm is governed by the mutually antagonistic effects of the transcription factors Oct4 and Cdx2. Evidence indicates that suppression of Oct4 expression in the trophectoderm is mediated by Cdx2. Nonetheless, the underlying epigenetic modifiers required for Cdx2-dependent repression of Oct4 are largely unknown. Here we show that the chromatin remodeling protein Brg1 is required for Cdx2-mediated repression of Oc ...[more]

PMID: 20485553

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Project description:In mouse blastocysts segregation of the inner cell mass (ICM) and the trophectoderm (TE) is regulated by the mutually antagonistic effects of the transcription factors Oct4 and Cdx2 expressed in the ICM and TE, respectively. In contrast, in other species such as bovine and human, Oct4 is not restricted to the ICM and continues to be expressed in the Cdx2-positive TE. A recent comparative study of the bovine and mouse Oct4 promoters revealed that additional mechanisms might act in conjunction with Cdx2 to downregulate Oct4 expression in the mouse TE lineage. For instance, the mouse Oct4 distal enhancer contains an AP-2? (Tcfap2c) binding motif that is absent in the bovine and human Oct4 distal enhancer. Nonetheless, the functional relevance of Tcfap2c in Oct4 repression during mouse preimplantation development was not tested. To elucidate the role of Tcfap2c in Oct4 expression an RNA interference approach was utilized. Depletion of Tcfap2c triggered a decrease in Oct4 expression at the 8-cell and morula stage. Remarkably, at the blastocyst stage depletion of Tcfap2c and/or its family member Tcfap2a had no effect on Oct4 repression. To test whether Tcfap2c interacts with Oct4 to positively regulate Oct4 expression, chromatin immunoprecipitation and in situ co-immunoprecipitation analyses were performed. These experiments revealed Tcfap2c and Oct4 binding were enriched at the Oct4 distal enhancer in embryonic stem (ES) cells, but were rapidly lost during differentiation into trophoblast-like cells when Oct4 became repressed. Moreover, Tcfap2c and Oct4 interactions were detected at the morula stage, but were lost during blastocyst formation. In summary, these data demonstrate that Tcfap2c is not required for Oct4 silencing in mouse blastocysts, but may be necessary for the maintenance of Oct4 expression during the 8 cell-to-morula transition. These findings support the notion Cdx2 is the predominant negative regulator of Oct4 expression during blastocyst formation in mice.

Dataset Information

Brg1 is required for Cdx2-mediated repression of Oct4 expression in mouse blastocysts.

Publications

Brg1 is required for Cdx2-mediated repression of Oct4 expression in mouse blastocysts.

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