Study On The Antioxidant Function Of Chi-miR-542-3p On Goat Caput Epididymal Epithelial Cells Based On The Whole Transcriptome

ceRNA is a competitive endogenous binding RNA.miRNA plays a role in the post-transcriptional regulation of m RNA through ce RNA mechanism.The existing researches on the regulatory role of non-coding RNA in goat caput epididymis are unclear.Therefore,the research on the regulatory mechanism of ce RNA in goat caput epididymis can lay a theoretical foundation for understanding goat caput epididymis development and sperm maturation in the epididymis.This study will use high-throughput sequencing technology,gene cloning technology,and bioinformatics methods to predict the differentially expressed ce RNAs regulatory mechanism in sequences and GPx5 gene’s ce RNA network in the glutathione pathway.The binding of chi-miR-542-3p with potential target genes GPx5,IDH1,and lnc RNA-MSTRG.57825.2 was verified by a dual fluorescent enzyme reporter system.Finally,effects of chi-miR-542-3p on the antioxidant function of in caput epididymal epithelial cells of Taihang goat were explored by overexpression and inhibition of chi-miR-542-3p.The results are as follows:1.The GPx5 gene of caput epididymis of Taihang goat was cloned.The total length of CDS was 660 bp,5’-UTR 59 bp,and 3’-UTR 690 bp.Based on the sequence of the GPx5 gene and whole transcriptome high-throughput sequencing,we constructed a ce RNA regulatory network associated with the caput epididymis and the caput epididymis glutathione pathway.The ce RNA network showed that GPx5,IDH1,and lnc RNA-MSTRG.57825.2 can bind chi-miR-542-3p in caput epididymis of Taihang goat.Finally,eight differentially expressed mRNA,lnc RNA,and miRNA in the tissues of young and adult Taihang goats were detected by q RT-PCR.The q RT-PCR results showed that the results of high-throughput sequencing were more reliabl.2 The results of dual fluorescent reporter assay showed that chi-miR-542-3p could significantly reduce the fluorescence intensity of dual fluorescent reporter vector containing GPx5-3′-UTR fragment,IDH1-3′-UTR fragment,and lnc RNA-MSTRG.57825.2fragment.The caput epididymal epithelial cells of Taihang goat were isolated and cultured in vitro.chi-miR-542-3p was overexpressed or silenced in the caput epididymal epithelial cells.It was found that after the overexpression of chi-miR-542-3p,the two potential target genes GPx5 and IDH1 were significantly down-regulated at the m RNA level,and the protein level of GPx5 was also considerably down-regulated,which reduced antioxidant capacity in the epididymal epithelial cells.The results of chi-miR-542-3p silencing showed the opposite results.In conclusion,a large number of the differentially expressed ce RNAs regulation mechanisms were found in caput epididymis between Taihang goat kids and adult Taihang goats.The preliminary results showed that chi-miR-542-3p can inhibit the expression of IDH1 and GPx5 genes through the ce RNA mechanism,regulating the antioxidant function of caput epididymal epithelial cells.The results provided a scientific basis for analyzing the mechanism of miRNA and lnc RNA on caput epididymal development and sperm maturation in Taihang goats.