猪GSKIP基因ceRNA调控网络及其蛋白互作分析

ceRNA regulatory network of GSKIP gene in pig and corresponding protein interaction analysis

利用RT-PCR技术从版纳微型猪近交系睾丸组织中扩增出GSKIP全长编码序列并分析其分子特征,对GSKIP基因进行功能注释,分析其与非编码RNA(miRNA和lnRNA)间的调控关系,并构建ceRNA转录调控网络,进一步分析GSKIP蛋白的基本功能,构建多物种进化树和相互作用蛋白网络.通过RT-PCR获得了BMI GSKIP CDS区全长420 bp,该基因含4个外显子;基因功能分析表明,GSKIP主要参与Wnt信号通路;基因靶向作用分析表明,猪GSKIP受ssc-miR-181c、ssc-miR-181b、ssc-miR-181a、ssc-miR-335和miR-644-5p等5个miRNA的靶向调控;蛋白质结构分析表明,GSKIP包含1个DUF727结构域,α螺旋在二级结构中占比最高;多物种进化分析表明,BMI GSKIP氨基酸序列在哺乳动物中较为保守,与绵羊和牛的亲缘关系最为接近;蛋白互作网络分析表明,BMI GSKIP与10个蛋白存在相互作用.

The full-length coding sequence of GSKIP was amplified from the testis tissues of Banna mini-pig inbred line(BMI) by reverse transcription PCR(RT-PCR) for its function annotation, regulatory relationship analysis with non-coding RNA(miRNA and lnRNA) and ceRNA transcriptional regulatory network analysis. Meanwhile, the basic function of GSKIP protein was analyzed, as well as multi-species evolutionary tree and protein-protein interaction network. The result of RT-PCR revealed 420 bp coding region and 4 exons in the GSKIP gene of BMI. Function and targeted gene analysis indicated that the GSKIP was mainly involved in Wnt signaling pathway, and was regulated by 5 miRNAs, including ssc-miR-181 c, ssc-miR-181 b, ssc-miR-181 a, ssc-miR-335 and miR-644-5 p. Protein GSKIP contained a DUF727 domain, and α-helix accounted for the highest proportion of secondary structure. Evolution relationship analysis revealed relatively conserved amino acid sequences of BMI GSKIP in mammals with the closest genetic relationship with sheep and cattle. Protein-protein interaction network analysis suggested that pig GSKIP interacted with 10 other proteins.