lncRNA-M7s1—miR-7通过Raf/MEK/ERK信号通路调控牛腺垂体FSH分泌机制的研究

The secretion of FSH by the adenohypophysis plays a crucial role in mammalian reproductive regulation. It is of great significance to determine the molecular mechanism of FSH secretion regulation for the artificial regulation of animal reproduction. Our previous study found that multiple miRNAs such as miR-7 can act on the GnRH signaling pathway and regulate the secretion of FSH in the bovine anterior pituitary cells. Combined with the sequencing results of the whole transcriptome of bovine adenohypophysis gland, several lncRNAs that may participate in the regulation were predicted. Based on the previous works, this project will reveal the molecular mechanism that miRNA-7 targets Raf gene and regulates the secretion of FSH. The site of interaction between lncRNA-M7s1 and miRNA-7 will be identified systematically. The mechanism of action of ceRNA will be analyzed. Further verify that lncRNA-M7s1 may directly target FSHβ gene. Finally, achieve the comprehensive analysis of lncRNA-M7s1 and miR-7 through the Raf/MEK/ERK signaling pathway regulating the secretion of FSH pituitary gland FSH molecular mechanism. The results will provide a theoretical basis for revealing the molecular mechanism of FSH secretion and provide theoretical support for improving the artificial regulation system of bovine reproduction.

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。课题组前期发现miR-7等多个miRNA能够作用于FSHβ基因，调控牛腺垂体细胞FSH的分泌；结合牛腺垂体全转录组测序结果，预测到了多个可能参与调控的lncRNA。本项目在前期工作的基础上，解析miRNA-7靶向作用于Raf基因，调控FSH分泌的分子机制，系统识别并鉴定LncRNA-M7s1与miRNA-7的相互作用位点，揭示其ceRNA作用机制，进一步验证lncRNA-M7s1直接靶向作用于FSHβ基因的可能，实现全面解析LncRNA-M7s1—miR-7通过Raf/MEK/ERK信号通路调控牛腺垂体FSH分泌的分子机制。研究结果将为揭示FSH分泌调控的分子机理提供理论基础，并为完善牛繁殖过程的人工调控体系提供理论支撑。

作为腺垂体合成并分泌的促性腺激素之一， FSH积极参与哺乳动物的繁殖调控。深入探究调控FSH合成与分泌的分子机制对丰富生殖内分泌学基础理论、实现动物繁殖的人工调控具有重要的借鉴意义。近年来，lncRNA已被发现在动物代谢活动、生殖发育、疾病发生等方面均发挥关键调控作用。然而， lncRNA在FSH合成与分泌过程中是否具有潜在的调控作用仍有待进一步探究。基于以上研究背景并结合前期研究基础，本研究成功预测到了能够与miR-433靶向作用的lncRNA-m433s1以及能够与miR-18a-5p靶向作用的lncRNA-m18as1。由于牛腺垂体组织供应受限，经比对保守性后在大鼠腺垂体原代细胞上进行机制验证。研究结果证实了以下两条调控FSH合成与分泌的ceRNA机制：（1）lncRNA-m433s1能够作为miR-433的分子海绵，通过行使ceRNA机制来降低miR-433对Fshβ的抑制作用，并进一步调节FSH的合成与分泌；（2）lncRNA-m18as1作为miR-18a-5p的分子海绵，竞争性结合miR-18a-5p调控Smad2的表达，进而通过SMAD2/3信号通路调控FSH的合成与分泌。为进一步挖掘调控哺乳动物FSH合成与分泌的分子调控机制，通过全转录组测序技术，对GnRH处理后表达显著差异的circRNA进行鉴定并成功构建了GnRH处理前后的差异circRNA表达谱，为进一步探索非编码RNA调控FSH合成与分泌的机制提出了新的研究思路。综上，本研究解析了“lncRNA-m433s1-miR-433-Fshβ”、“lncRNA-m18as1 miR-18a-5p-SMAD2/3信号通路-Fshβ”两条分子机制，为揭示FSH合成与分泌的分子机理提供了理论基础，并为完善牛繁殖过程的人工调控体系提供了理论支持。