名贵经济鱼类-鳜鱼肌肉分化生长调控分子机理研究

Molecular mechanisms of muscle differentiation and growth in Chinese mandarin fish.The molecular regulation of fish muscle development and growth is one of the most important issues in fish muscle biology. In this proposal, we plan to use the economically important fish species, Chinese mandarin fish as a model system to address the major scientific issues regarding the origin and cellular characteristics of muscle stem cells (satellite cells), the mechanism of satellite cell activation, proliferation and self-renewal during muscle development and growth. The project will be specifically focused on the following aims: 1)to study the histone methyltransferase SmyD1 and transcription factors Pax3, Pax7, Myf5 and MyoD regulatory networks in controlling muscle satellite cell proliferation and self-renewal;2)to characterize the formation of slow and fast muscles via quantitative, and qualitative analysis of coordinated expression of muscle-specific transcription factors and structural genes；3)to isolate and determine the expression profile of microRNAs in skeletal muscles of mandarin fish, and to determine their biological functions in satellite cell activation, proliferation and self-renewal during muscle development and growth; 4)to establish a mandarin fish model to study the effect of embryonic temperature control and diet compensatory stimulation on fish muscle growth. Overall, above studies address the fundamental questions in the cellular and molecular regulation of fish muscle development and growth using diverse approaches encompassing cellular and molecular biology,developmental biology and biochemical analyses. Collectively,these studies will provide significant contribution to the current understanding of muscle development and growth in Chinese mandarin fish. More importantly, these findings also provide the technical implications for aquacultural application.

鱼类肌肉分化生长及其调控机理一直是鱼类发育分子生物学领域研究的热点之一。本项目以名贵经济鱼类-鳜鱼为研究对象，拟阐明其肌卫星细胞起源、定向分化以及肌肉生长发育的生物学特征和调控机理。该项目将聚焦：1)阐明组蛋白甲基化酶SmyD1、转录因子Pax3和Pax7、成肌因子Myf5和MyoD的表达调控网络，揭示肌卫星细胞激活、增殖和分化的分子机理；2)阐明幼鱼和成鱼肌肉卫星细胞的再激活、分化和迁移与肌肉生长的关系；3)筛选与肌卫星细胞激活、分化和肌肉生长特异miRNA分子并解析其功能和调控作用；4)建立温度控制和补偿生长养殖模式,解析环境因子对肌肉生长影响的机理。因此,本项目从组织细胞学和发育分子生物学角度全面深入揭示鳜鱼肌肉分化生长特征和调控机理，具有重要鱼类发育生物学理论意义和应用前景。

鳜鱼因其味道鲜美，肉质细嫩，营养价值高，深受广大消费者欢迎，已成为我国大力推广的重要养殖对象。本项目采用综合细胞学和分子生物学技术对鳜鱼肌细胞，肌纤维分化以及营养调控开展了系统研究。采用细胞切片和肌卫星标记鉴定了早期肌原细胞起源于原肠期胚胎并在幼鱼或成鱼各生长期均有肌卫星细胞存在，揭示了鱼类终生生长的原理；首次克隆了影响鳜鱼肌纤维分化的关键基因蛋白甲基化酶Smyd1a和Smyd1b，并分析了该基因的功能和调控机制；分别克隆了系列与肌肉反应分化相关的结构基因和生肌调节因子或转录因子，包括Pax3 、Pax7、Myf5、 和MyoD等，并对该类基因的在鳜鱼肌肉组织中时空表达规律和调控网络进行了分析研究，揭示肌卫星细胞激活、增殖和肌肉分化生长的分子机理；通过肌细胞的在不同生长阶段的计量分析，阐明幼鱼和成鱼肌肉细胞的增殖和增大与肌肉生长的内在关系；采用高通量RNA测序（RNAseq）和基因芯片技术比较分析了鳜鱼两种类型肌肉组织的蛋白质和microRNA的表达谱特征，鉴定了多个影响鳜鱼肌肉分化生长的特异microRNA分子，预测和分析了该特异microRNA分子的靶基因和调控机制，阐明了microRNA 与SMYD1基因调控肌肉分化的机制和网络；为提高鳜鱼的养殖效益，我们建立了以饥饿和再投喂的养殖模式，即补偿生长养殖,并解析了补偿生长影响鳜鱼生长的基因包括miroRNA分子的表达和调控肌肉生长影响的分子机理。概括起来,本项目从组织细胞学和发育分子生物学角度深入揭示了鳜鱼肌肉分化生长特征和调控机理，具有重要鱼类发育生物学理论意义和指导鳜鱼生产应用价值。本项目相关结果，发表研究论文27篇，其中在Scientific Reports、BMC genetics, PLoS ONE 和Molecular Biology of the Cell等杂志发表SCI收录论文20篇，获国家授权发明专利2项，出版著作2部，制定湖南省省级标准1项，获湖南省自然科学奖三等奖1项。