Transcription factor TFAP2C affects porcine early embryo development via regulating epigenetic modification

Abstract

AbstractTranscription factors (TFs) have the potential function in regulating gene expression. Transcription factor TFAP2C plays important roles in the regulation of post-implantation embryonic development in mice, the reprogramming process, trophectoderm formation and carcinogenesis, but its role in porcine early embryo development remains unclear. This study was conducted to investigate the role of TFAP2C in porcine early embryo development using siRNA cytoplasmic injection. The RNAseq and immunofluorescence staining were performed to detect gene expression, and ChIP and dual luciferase reporter assays were used to elucidate the mechanism. The results showed that the deficiency of TFAP2C could lead to embryonic development disorder. The percentage of the blastocyst in theTFAP2Cknockdown (TFAP2C-KD) group (7.76±1.86%) was significantly decreased compared to the control group (22.92±1.97%) (P**<0.01). The RNAseq results showed that 1208 genes were downregulated and 792 genes were upregulated after siRNA injection. The expression of epigenetic modification enzymes KDM5B, SETD2 (P**<0.01)etc. were significantly elevated inTFAP2C-KDgroup. Meanwhile, the modification levels of H3K4me3, H3K4me2 and H3K9me3 (P*<0.05) were significantly decreased, and the modification levels of H3K36me3 (P**<0.01) and DNA methylation (P**<0.01) were significantly increased inTFAP2C-KD group. DNMT1 was mostly expressed in cytoplasm in the control group, while it was mainly expressed in nuclei in theTFAP2C-KD group. In addition, TFAP2C could bind to the promoter region ofSETD2, and the mutation of the TFAP2C binding site resulted in increased activity ofSETD2promoter (P**<0.01). The knockdown of TFAP2C affects histone modification and DNA methylation by regulating the expression ofSETD2, KDM5B etc. and other genes, thereby inhibiting embryonic development. TFAP2C binds to the promoter region ofSETD2and acts as a hindrance protein. This study fills in the deficiency of TFAP2C in porcine early embryo development and provides theoretical support for animal husbandry production and biomedicine.Author SummaryThe correct activation of embryonic genes is required during early embryonic development, and the activation of these genes is subject to strict epigenetic regulation, such as DNA methylation, histone acetylation and methylation, with abnormalities in either leading to birth defects and developmental defects in individuals. TFs have specific binding motifs that regulate gene expression by binding to them. TFAP2C has been studied in post-implantation embryonic development and trophectoderm generation, however, the effect on early embryo development is unknown. Our findings suggest that TFAP2C deficiency disrupts gene expression patterns and leads to abnormal epigenetic modifications, resulting in abnormal embryo development. Furthermore, we found for the first time that TFAP2C can bind to the promoter region ofSETD2, thereby affecting early embryo development in pigs. This indicates the critical role of TFAP2C in early embryo development in pigs on one hand, and also provides theoretical support for livestock production and biomedicine.