Reciprocal regulation of TEAD4 and CCN2 for the trophectoderm development of the bovine blastocyst

Abstract

The first segregation at the blastocyst stage is the symmetry-breaking event to characterize two cell components; namely, inner cell mass (ICM) and trophectoderm (TE). TEA domain transcription factor 4 (TEAD4) is a well-known regulator to determine TE properties of blastomeres in rodent models. However, the roles of bovine TEAD4 in blastocyst development have been unclear. We here aimed to clarify the mechanisms underlining TE characterization by TEAD4 in bovine blastocysts. We first found that theTEAD4mRNA expression level was greater in TE than in ICM, which was further supported by TEAD4 immunofluorescent staining. Subsequently, we examined the expression patterns of TE-expressed genes;CDX2,GATA2andCCN2, in theTEAD4-knockdown (KD) blastocysts. These expression levels significantly decreased in theTEAD4KD blastocysts compared with controls. Of these downregulated genes, theCCN2expression level decreased the most. We further analyzed the expression levels of TE-expressed genes;CDX2,GATA2andTEAD4in theCCN2KD blastocysts. Strikingly, theCCN2KD blastocysts showed the downregulation ofCDX2,GATA2andTEAD4. Furthermore, the ratio of TE-to-ICM cell numbers in theCCN2KD blastocysts significantly decreased compared to controls. To our knowledge, this is the first study showing the regulation ofCCN2expression thoroughTEAD4in mammalian embryos. Not only that, this study also provides evidence that reciprocal regulation ofTEAD4andCCN2is required for TE development with appropriate gene expression in bovine blastocysts.