miR-181家族对牛FSHβ基因转录后调控及FSH分泌作用机制的研究

The hormones secreted by pituitary play a major role in the regulation of animal’s reproductive process, and miRNAs exert effects on the secretion of hormones in pituitary by gene post-transcriptional regulation. Our previous study suggested that there are significant differences of miRNAs’ expression between immature and sexually matured bovine pituitary.In order to validate the effect of the differentially express miRNA-181 family on FSHβ gene post-transcriptional regulation, we are going to using bioinformatics method to predict the target genes of the differentially expressed miRNAs and screening the miRNA-181 family which have the directly function about FSHβ genes expression. Then we will verify the relationship between miRNA-181 family and FSHβ gene post-transcriptional regulation in vitro by Dual-Luciferase Reporter Assay System. Furthermore, the transfect bovine pituitary cell with the miRNA-181 family and inhibitors in order to authenticate the influence on secretion the FSH and pituitary cell’s development.The results will clear miRNA-181 family which exert the function of post-transcription regulation about FSHβ, the results also reveals gene expression, regulation and molecular mechanisms of bovine pituitary gonadotropins.

哺乳动物腺垂体所分泌的FSH在繁殖过程中发挥着重要的调节作用，深入研究FSH合成与分泌的分子调节机制，有助于实现对动物繁殖过程的人工调控。申请人前期研究表明，在初情期前和性成熟的牛腺垂体中miR-181家族成员表达量差异显著，并且在大鼠腺垂体细胞中证实FSHβ基因是其靶基因。为了识别与注释miR-181家族成员对FSHβ基因转录后调控的影响，本研究拟采用生物信息学方法分析并验证miR-181家族成员对靶基因FSHβ的作用位点及其调控网络，筛选候选miRNA，构建候选miR-181 的agomir和antagomir并转染牛腺垂体细胞，验证其对牛腺垂体细胞增殖、凋亡及FSH分泌的影响；并分析长、短和超短反馈对候选miRNA及FSHβ基因表达的影响。研究结果将解析miR-181家族对牛FSHβ基因转录后调控的分子机制，并明确其对FSH分泌的影响，为家畜繁殖调控提供理论基础。

腺垂体所分泌的FSH在哺乳动物繁殖过程中起着至关重要的作用，明确FSH分泌调控的分子机制对实现动物繁殖过程的人工调控具有借鉴意义。本研究在前期工作的基础上，分析并验证了miR-181b对靶基因FSHβ作用位点及其调控网络，通过mimics和inhibitors转染牛腺垂体原代细胞，验证其对牛腺垂体原代细胞增殖、凋亡及FSH分泌的影响；并阐明了GnRH对候选miRNA及FSHβ基因表达的影响。另外还筛选出18个可能调控腺垂体FSHβ基因的差异表达的miRNA，并探明了miR-21-3p、miR-433、miR-186及miR-7a对腺垂体FSH分泌影响的机制。构建了性成熟前后牛及大鼠的腺垂体lncRNA表达谱，并进行了生物信息学分析, 探索lncRNA和miRNA对腺垂体分泌机制的调控作用。以上研究明确了对FSHβ基因具有转录后调控功能的miRNA，揭示了腺垂体分泌的FSH的表达调控规律与分子机制，将为miRNA应用于动物繁殖调控探索新的思路。