Molecular mechanisms of muscle stem cells transitioning intoquiescence

肌肉干细胞转变为静止状态的分子机制

• 批准号: 8538840
• 负责人: Christoph Lepper
• 金额: $ 37.83万
• 依托单位: CARNEGIE INSTITUTION OF WASHINGTON, D.C.
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-20 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8538840
• 关键词: ARHGEF5 gene; Accidents; Acute; Adult; Alleles; Automobile Driving; Behavior; Bioinformatics; Biological Assay; Candidate Disease Gene; Cell Adhesion; Cell Death; Cell Polarity; Cell Proliferation; Cell Separation; Cell Therapy; Cells; Characteristics; Chimeric Proteins; Clinical; Cytoskeleton; Data; Defect; Development; Epigenetic Process; Essential Genes; Estrogen Receptors; Future; Gene Expression Profile; Genes; Genetic; Injury; Intramuscular; Knock-in Mouse; Life; Locomotion; Longitudinal Studies; Mechanics; Mediating; Messenger RNA; Modeling; Molecular; Movement; Mus; Muscle; Muscle satellite cell; Muscular Dystrophies; Natural regeneration; Newborn Infant; Pathway interactions; Play; Population; Posture; Process; Production; Proliferating; Property; Proteins; Regulation; Reporter; Role; Signal Transduction; Skeletal Muscle; Sorting - Cell Movement; Source; Staging; Stem cells; Tamoxifen; Testing; Time; Transplantation; Viral; base; combat; comparative; critical period; gene function; knock-down; mouse model; muscle regeneration; mutant; novel; programs; recombinase; repaired; satellite cell; screening; small hairpin RNA; stem cell population; transcription factor

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.

项目总结 哺乳动物的肌肉系统为运动提供机械力。肌肉显示出非凡的能力 从肌肉干细胞储存库中再生。肌肉干细胞如何从活跃的增殖转化为 成年期为肌肉修复/再生的储备种群 一生都是未知的。我获得的证据表明，在小鼠模型中，这一过程取决于 Pax7基因的功能，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)转录组在发育到成人的转变过程中的变化，3)其他 调节这一转变的基本基因(与Pax7平行或下游)。 目的1：检测突变型Pax7卫星细胞的细胞和分子缺陷。何时以及如何实现 PAX7突变细胞在发育到成体的转变过程中会出现缺陷？我会结合过多的化验 用于确定几种肌肉干细胞特性，包括肌肉分化、细胞增殖、细胞 死亡、利基占有率和细胞极性。这些分析将与时间进程研究一起进行，从 从新生儿到成人。 目的2：对Pax7对照和突变型卫星细胞进行mRNA序列分析。转录组是什么？ 在此过渡过程中发生的变化以及Pax7管理的具体流程是什么？比较时间 对照细胞和Pax7突变细胞从发育到成体转录本的过程研究 过渡期将使用mRNA-SEQ进行，然后进行生物信息学分析。 目的3：筛选卫星细胞功能的候选基因。基因是什么(除了 PAX7)对于肌肉干细胞从发育到成体的转变至关重要？从目标2中，我将选择最多50个基因 测试它们在这方面的功能；它们可以独立运行，也可以在Pax7的下游运行。我会发展的 一种结合基因敲除和肌内注射快速筛选这些基因功能的新方法 肌肉干细胞移植。