The Role of MicroRNAs in Vertebrate Muscle Development.

描述(申请人提供)：这个项目的目标是确定肌肉特异的microRNAs(MiRNA)在体内调节非洲爪蛙体节和肌肉形态发生的作用。MiRNAs是一种小的、非编码的RNA序列，通过结合特定的信使RNA并阻止其翻译，在转录后调节基因。它们已被证明在胚胎发育和疾病中发挥着重要作用。特别令人感兴趣的是三个肌肉特异的RNA，miR-1，miR-206和miR-133，它们在心肌和骨骼肌组织的形成和维持中发挥重要作用。这些miRNAs与几种不同的肌肉相关疾病有关。尽管人们对这些miRNAs在发育和疾病中的作用越来越感兴趣，但到目前为止还很少进行体内研究。因此，我们的实验室已经开始使用非洲爪哇作为我们的模型系统来研究miR-206的作用。我们的初步研究表明，miR-206在轴旁中胚层中表达，在正确的时间和地点介导肌肉发育。此外，miR-206的表达下调会导致肌细胞的分割、体节形成以及肌细胞的伸长和排列出现明显的缺陷。鉴于每个miRNA被预测为调节不同的靶点，我们假设这三个miRNAs在调节胚胎体节和肌肉形态发生的细胞行为方面具有独特和互补的作用。我们将使用细胞和分子相结合的方法来检验这一假说：击倒每个miRNA对躯体和肌肉形态发生的影响(目标1)；过度表达每个miRNA对躯体和肌肉形成背后的形态发生运动的影响(目标2)；以及miR-1、miR-206和miR-133对形态发生的组合作用(目标3)。总的来说，这项工作将在体内全面地研究这三个miRNAs在胚胎发生过程中对体节和肌肉形成的作用，并将识别在这一形态过程中重要的细胞机制，这些机制由这些miRNAs调控。该SC-3项目将通过与加州大学伯克利分校哈兰德实验室的持续合作，进一步推动PI的开发。哈兰德实验室是非洲爪哇一流的实验室，开发了许多尖端工具，用于研究非洲爪哇胚胎发育期间的基因调控。在这种合作努力和所要求的资金的帮助下，PI将提高她的研究生产力，以便过渡到主流资金。该项目还将为全国文化最多元化的综合性公立大学之一的大量本科生和硕士生提供一个重要研究领域的研究培训机会，这些学生主要是代表不足的少数族裔。

DESCRIPTION (provided by applicant): The goal of this project is to determine the role that muscle-specific microRNAs (miRNA) play in vivo to regulate somite and muscle morphogenesis in the frog, Xenopus laevis. miRNAs are small, non-coding sequences of RNA that regulate genes post-transcriptionally by binding specific messenger RNA and blocking its translation. They have been shown to play a major role in embryogenesis and diseases. Of particular interest are three muscle-specific RNAs, miR-1, miR-206 and miR-133, which play an important role in the formation and maintenance of cardiac and skeletal muscle tissue. These miRNAs have been linked to several different muscle-related diseases. Although there is growing interest in the role of these miRNAs in development and disease, very few in-vivo studies have been conducted to date. Thus, our lab has begun to examine the role of miR-206 using Xenopus laevis as our model system. Our preliminary studies indicate that miR-206 is expressed in the paraxial mesoderm at the right time and place to mediate muscle development. Further, knockdown of miR-206 expression leads to significant defects in segmentation, somite formation and muscle cell elongation and alignment. Given that each miRNA is predicted to regulate different targets, we hypothesize that these three miRNAs have both unique and complementary roles in regulating the cell behaviors that underlie embryonic somite and muscle morphogenesis. We will test this hypothesis using a combination of cell and molecular approaches to examine: the impact of knocking down each miRNA on somite and muscle morphogenesis (Aim 1); the impact of overexpressing each miRNA on the morphogenetic movements underlying somite and muscle formation (Aim 2); and the combinatorial roles of miR-1, miR-206 and miR-133 on morphogenesis (Aim 3). Collectively, this work will provide a comprehensive in vivo examination of the role of these three miRNAs on somite and muscle formation during embryogenesis and will also identify cellular mechanisms that are important in this morphological process that are regulated by these miRNAs. This SC-3 project will further the PI's development through ongoing collaboration with the Harland laboratory at UC Berkeley, a premier Xenopus lab that has developed many cutting edge tools for investigating gene regulation during Xenopus embryonic development. With the help of this collaborative effort and the requested funding, the PI will increase her research productivity in order to transition to mainstream funding. The project will also provide research training opportunities in an important field of study for large numbers of undergraduate and master's students, primarily underrepresented minorities, at one of the nation's most culturally diverse comprehensive public universities.