LncRNA通过激活TGFβ/SMAD2信号通路调控湖羊垂体促性腺激素分泌的机制研究

垂体促性腺激素(FSH和LH)是调控家畜繁殖力的关键性激素。申请人前期对高低繁湖羊垂体测序，筛选到一个新的差异表达lncRNA SM2及其靶基因SMAD2，且其与促性腺激素的分泌存在相关性，推测lncRNA SM2可能通过靶向TGFβ/SMAD2通路调控促性腺激素分泌，但其机制尚不清楚。因此，本项目拟首先利用RACE、FISH、核质分离等技术研究lncRNA SM2的表达特征、全长序列信息及亚细胞定位；然后通过CRISPRi/a等技术明确lncRNA SM2调控促性腺激素分泌的作用及其与SMAD2的靶向关系；再综合采用RNA pull down、RIP等技术解析lncRNA SM2的具体作用机制；最后通过构建绵羊ES来源的垂体类器官模型以及小鼠活体模型，系统阐明lncRNA SM2参与调控垂体促性腺激素分泌的作用和调控网络，为绵羊多胎的分子调控机制和生殖调控手段的开发提供新的思路。

Pituitary gonadotropins (including FSH and LH) are the key hormones regulating the fecundity of domestic animals. In our previous study, a new lncRNA SM2 and its target gene SMAD2 were screened in Hu sheep pituitary associated with prolificacy, which were also related to the secretion of gonadotropin. We speculate lncRNA SM2 may regulate gonadotropin secretion of Hu sheep by targeting TGFβ/Smad2 pathway, but the mechanism remains unknown. Thus, in this study, we will identify its expression characteristics, full-length sequence information and subcellular localization using RACE, FISH, and nuclear/cytoplasmic fractionation ; then we will study the effect of lncRNA SM2 on gonadotropins secretion and its regulation of TGFβ/Smad2 signal pathway through crispri/a methods, and clarify the action mechanism of lncRNA SM2 by RNA pull down, RIP. Finally we will explore the effect of lncRNA SM2 involved in the regulation of gonadotropin secretion by sheep pituitary organoids model and in vivo mouse model. This project will provide new ideas for the regulation mechanism of sheep prolificacy and animal reproductive regulation.