微肽蛋白UBL5通过Ube2g1可变剪接调控牛胚胎基因组激活的分子机制

可变剪接是调节基因表达和产生蛋白质组多样性的重要机制之一。我们前期发现具有可变剪接调控功能的微肽蛋白UBL5在牛胚胎基因组激活期表达量显著升高，干扰UBL5会降低牛克隆胚胎囊胚率。同时发现E2泛素结合酶Ube2g1基因发生可变剪切模式改变，Ube2g1内含子中具有UBL5依赖性的5’剪接位点。但是，微肽蛋白UBL5是否通过调控Ube2g1可变剪接并进一步影响牛早期胚胎发育尚不清楚。本研究拟以牛8-16细胞期克隆胚胎为研究对象，利用过表达和干扰技术验证UBL5对Ube2g1基因可变剪接的调控；构建Ube2g1选择性表达载体，转染Ube2g1-/-细胞,获得无内源性Ube2g1影响的转基因细胞株，核移植后分析Ube2g1不同转录本编码蛋白对牛早期胚胎发育的影响。本研究对于揭示微肽蛋白UBL5通过可变剪接调控牛早期胚胎发育的分子机制提供理论依据，为突破体外培养胚胎发育阻滞方面的研究提供新的思路。

Alternative splicing(AS) of pre-mRNA is a key component in the regulation of gene expression in mammals and plays an important role in many biological processes. We recently found that UBL5, which affects AS through UBL5-SART1 interaction, was significantly up-expressed in bovine SCNT embryos during embryonic genome activation (EGA) stage. The AS patterns of Ube2g1, a key component of the ubiquitin-binding enzyme E2, had changed during this stage, and the UBL5 dependent 5’splice site (GUGAGU) also found in this gene. But, it is unclear whether UBL5 regulates Ube2g1 alternative splicing and further affects the early development of bovine embryos. In order to validate this hypothesis, this project was designed to study the effect of micropeptide UBL5 on the AS events of Ube2g1 gene by interfering with and overexpressing UBL5. The effects of different spliceosome of Ube2g1 gene on early embryonic development will be analyzed by constructing Ube2g1 gene selective expression vector. This research may provide new insights into revealing the mechanisms of micropeptide protein UBL5 in mammalian embryonic genome activation, and play an important role on bovine preimplantation embryos to overcome developmental block in vitro.