Molecular mechanisms of muscle stem cells transitioning intoquiescence

项目摘要 哺乳动物的肌肉组织为运动提供机械力。肌肉显示出非凡的能力， 从肌肉干细胞库中再生。肌肉干细胞是如何从一个活跃增殖的 发育期间的种群转变为成年期肌肉修复/再生的储备种群 一生都不知道。我已经获得的证据表明，在小鼠模型中，这一过程取决于 Pax7基因的功能，它编码一种转录因子。我计划以下目标来定义1） Pax7的确切作用，2）发育到成年过渡期间的转录组变化，以及3）其他 介导这种转变的必需基因（与Pax7平行或下游作用）。 目的1：确定Pax7突变卫星细胞的细胞和分子缺陷。何时以及如何 Pax7突变细胞在发育到成人的过渡中变得有缺陷？我会联合收割机 用于确定肌肉干细胞的几种特性，包括肌肉分化、细胞增殖、细胞分化、细胞增殖、细胞周期等。 死亡、生态位占有和细胞极性。这些试验将通过时间过程研究进行， 从新生儿到成年人 目的2：进行Pax7对照和Pax7突变卫星细胞的mRNA-seq。什么是转录组 在此过渡期间发生了哪些变化，Pax7管理的具体流程是什么？比较时间 在整个发育到成人的过程中，对照和Pax7突变细胞之间的转录组的课程研究 过渡期将使用mRNA-seq进行，然后进行生物信息学分析。 目的3：筛选卫星细胞中的候选基因。什么是基因（除了 Pax7）对于发育到成人肌肉干细胞的转变至关重要？从目标2中，我将选择多达50个基因， 以测试它们在这方面的功能;它们可以独立地或在Pax7下游起作用。I will develop 通过基因敲除和肌肉注射相结合快速筛选这些基因功能的新方法 肌肉干细胞移植

PROJECT SUMMARY The mammalian musculature provides mechanical force for locomotion. Muscles display remarkable ability to regenerate from reservoirs of muscle stem cells. How muscle stem cells transition from an actively proliferating population during development into a reserved population in adulthood for muscle repair/regeneration throughout lifetime is not known. I have obtained evidence that, in the mouse model, this process depends on the function of the Pax7 gene, which encodes a transcription factor. I plan the following aims to define 1) the precise role of Pax7, 2) the transcriptome changes during the development-to-adult transition, and 3) other essential genes (acting in parallel or downstream of Pax7) that mediate this transition. Aim 1: Determining the cellular and molecular defects of Pax7 mutant satellite cells. When and how do Pax7 mutant cells become defective in the development-to-adult transition? I will combine a plethora of assays for determining several muscle stem cell properties, including muscle differentiation, cell proliferation, cell death, niche occupancy, and cell polarity. These assays will be conducted with time course studies from newborn to adult. Aim 2: Performing mRNA-seq of Pax7 control and Pax7 mutant satellite cells. What are the transcriptome changes during this transition and what are the specific processes governed by Pax7? A comparative time course study of transcriptomes between control and Pax7 mutant cells throughout the development-to-adult transitional period will be performed using mRNA-seq, followed by bioinformatic analysis. Aim 3: Screening candidate genes for their function in satellite cells. What are the genes (in addition to Pax7) critical for the development-to-adult muscle stem cell transition? From Aim 2, I will select up to 50 genes to test for their function in this aspect; they may either act independently or downstream of Pax7. I will develop a novel assay for rapid screening of these genes' functions by combining gene knock-down and intramuscular transplantation of muscle stem cells.