Dataset Information

Knockdown of p66Shc Alters Lineage-Associated Transcription Factor Expression in Mouse Blastocysts.

ABSTRACT: The p66Shc adaptor protein regulates apoptosis and senescence during early mammalian development. However, p66Shc expression during mouse preimplantation development is upregulated at the blastocyst stage. Our objective was to determine the biological function of p66Shc during mouse blastocyst development. In this study, we demonstrate that a reduced p66Shc transcript abundance following its short interfering RNA (siRNA)-mediated knockdown alters the spatiotemporal expression of cell lineage-associated transcription factors in the inner cell mass (ICM) of the mouse blastocyst. P66Shc knockdown blastocysts restrict OCT3/4 earlier to the inner cells of the early blastocyst and have ICMs containing significantly higher OCT3/4 levels, more GATA4-positive cells, and fewer NANOG-positive cells. P66Shc knockdown blastocysts also show a significantly reduced ability to form ICM-derived outgrowths when explanted in vitro. The increase in cells expressing primitive endoderm markers may be due to increased ERK1/2 activity, as it is reversed by ERK1/2 inhibition. These results suggest that p66Shc may regulate the relative abundance and timing of lineage-associated transcription factor expression in the blastocyst ICM.

SUBMITTER: Edwards NA

PROVIDER: S-EPMC6209429 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Knockdown of p66Shc Alters Lineage-Associated Transcription Factor Expression in Mouse Blastocysts.

Edwards Nicole A NA Watson Andrew J AJ Betts Dean H DH

Stem cells and development 20180906 21

The p66Shc adaptor protein regulates apoptosis and senescence during early mammalian development. However, p66Shc expression during mouse preimplantation development is upregulated at the blastocyst stage. Our objective was to determine the biological function of p66Shc during mouse blastocyst development. In this study, we demonstrate that a reduced p66Shc transcript abundance following its short interfering RNA (siRNA)-mediated knockdown alters the spatiotemporal expression of cell lineage-ass ...[more]

PMID: 30091687

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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

Knockdown of p66Shc Alters Lineage-Associated Transcription Factor Expression in Mouse Blastocysts.

Publications

Knockdown of p66Shc Alters Lineage-Associated Transcription Factor Expression in Mouse Blastocysts.

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