miR-34b/c在鲟鱼性别分化中的调控作用及其分子机制

鱼类性腺发育和性别分化受下丘脑-垂体-性腺轴（HPG）的调控。目前鲟鱼养殖产业急需性别调控技术，但阐明鲟鱼性别决定与分化机制是首要解决的关键科学问题。新近研究表明microRNA介导靶基因的转录后表达是动物性别决定与分化的重要调控机制。我们首次报道了鲟鱼物种特异microRNA，从幼龄鲟鱼microRNA的性二态表达特性和功能分析中发现microRNA参与其性别分化调控，并鉴定miR-34b/c是鲟鱼性别分化中的重要候选基因。本项目拟以施氏鲟早期性别分化事件为研究主线，以miR-34b/c为切入点，采用SCP3免疫组化等技术揭示其性腺早期分化特征，明确miR-34b/c在脑和性腺的时空表达谱；利用microRNA过表达/干扰技术从体外到体内深入研究miR-34b/c对鲟鱼性别分化的影响，并解析其作用的分子机制，将为鲟鱼养殖产业性别调控提供新的基因资源和技术依据，亦可丰富鱼类生殖调控理论。

Fish gonadal sex differentiation is regulated by the hypothalamo-pituitary-gonad (HPG) axis. Sturgeon aquaculture needs more female populations and reproduction-related regulation techniques, but the mechanism of sex determination and differentiation in sturgeon is still unclear. Recent studies indicate that post-transcriptional regulation of sex differentiation related genes expression mediated by microRNAs (miRNAs) is the important regulatory factors in animals. In previous studies, we have found many sturgeon miRNAs with sex-dimorphic expression pattern between ovary and testis by high-through sequencing and microarray. We concluded that miRNAs may also play a key role in sex differentiation regulation of sturgeonfrom functional prediction.Among them, miR-34b/c were the especially important candidate genes for sex differentiation, which may regulate the biological process by targeted AR gene expression. Therefore, in this study, based on our previous studies in Amur sturgeon (Acipenser schrenckii), which is an important economic sturgeon species in China, we plan to find the spatial-temporal expression profile of miR-34b/c in brain and gonad tissues during early sex differentiation, and characterizations of early differentiation of gonad by immunohistochemistry using SCP3 antibody. Then, comprehensive regulation relationships will be verified by luciferase assays at the cellular level between miR-34b/c and 3’UTR of targeted AR gene. Moreover, miR-34b/c overexpression and inhibition experiment in vitro and in vivo will be carried out to explore its effect on sex differentiation, respectively. Our research will clarify regulation roles of miR-34b/c by targeted AR gene expression in early sex differentiation process and expand the molecular mechanism of sex determination and differentiation regulated by miRNAs in sturgeon. The results of miRNAs involved in sex determination and differentiation can provide us with novel genes and techniques for sturgeon breeding, and enrich the theory of fish reproductive regulation.