睾丸优势表达蛋白RPL39L在小鼠精子释放过程中的作用和机制研究

Spermiation is an important biological event during spermatogenesis, it needs CLATHRIN-dependent TBCs formation, which leads to ES disaggregation, but its inherent mechanism is still unclear. We discovered a series of ribosome related proteins, which could bind specific sequences and promote the protein translation. Among them, here we focus on one protein named RPL39L, which is functional unknown. It is predominantly expressed in mice testis, RPL39L deficiency caused male infertility, but not in female. We demonstrated the spermiation failure in knockout mice due to the obstacle of ES disaggregation. We also found the CLATHRIN expression was significantly lower in knockout mice. The results of bioinformatics analysis suggested RPL39L might enhance the CLATHRIN translation through binding its specific region of RNA sequence. In this study, we will verify the spermiation failure in RPL39L deficiency mice through antibody localization and phenotypic assay. We will also try to elucidate the regulation role of RPL39L through binding clathrin RNA. This study is innovative and will provide a new theoretical basis for diagnosis and treatment of male infertility, which has important significance and clinical application value.

精子释放是精子发生过程中一个重要生物学事件，该过程依赖CLATHRIN装配TBCs，促进ES解聚，其启动机制尚不清楚。申请人前期在人睾丸蛋白质组谱系中筛选到一类核糖体相关蛋白，它们可以结合特定序列并促进下游蛋白翻译。其中我们关注到一个功能未知的蛋白RPL39L，其在小鼠睾丸中优势表达，缺失RPL39L的小鼠表现为雄性不育，雌性可育。睾丸切片染色结果显示精子释放障碍，且与ES解聚障碍有关，进一步研究发现ES解聚障碍与TBCs募集装配因子CLATHRIN的下调有关。生物信息学分析提示RPL39L蛋白可以结合clathrin RNA促进其翻译。本课题拟通过RPL39L定位研究和敲除小鼠表型分析深入揭示RPL39L缺失影响精子释放的具体作用；验证RPL39L结合clathrin RNA的促翻译作用，全面阐明RPL39L在精子释放过程中的作用。本研究为男性不育的诊疗提供了理论依据，具有重要临床意义。

本项目首先通过构建rpl39l基因敲除小鼠，分析了rpl39l基因缺失后的雄性亚生育的表型，其次通过睾丸和精子活体分析寻找相关亚生育表型的原因，我们结合HE染色和免疫荧光实验阐明了rpl39l缺失后精子释放障碍的机理，再次，我们又通过蛋白质组学研究探讨了精子释放障碍背后可能的分子机制，并且通过基因注释分析探讨了rpl39l在雄性生殖系统中可能的分子通路，本项目的研究为雄性不育的诊疗提供了理论依据和新的线索；为男性避孕药物的研发提供了新的靶点。