Muscle clock and weakness: diversity supplement

摘要：本项目的目的是为西尔瓦那提供研究和职业发展培训。 西德霍姆目前是佛罗里达大学的博士生。这份行政副刊是对PA的回应- 21-071：《促进健康相关研究多样性的研究补充资料(行政补充)》将促进 为候选人的职业发展，以及为母项目的使命做出贡献-定义 生物钟下游的分子靶标，调节肌肉结构，暗示 力量。 Sidhom女士提议的研究的主要目标将是开展与目标2相关的实验 家长拨款。首先，她将学习使用共聚焦成像方法和肌瘤抗体来确定 对照和肌肉特异性肌节的M线和Z线结构元素的变化 BMal1KO小鼠。其次，她将使用AAV方法恢复选定的肉瘤基因的表达， 例如TCAP，以测试这是否足以恢复结构和肌肉力量。Sidhom女士将积极工作 与科林·道格拉斯，另一名正在为家长提供助学金的博士生以及克里斯托弗·沃尔夫博士一起， 这个项目的博士后助理。埃瑟博士是R01的私家侦探，他将是西德霍姆的主要导师 在佛罗里达大学医学院。我们已经成立了她的委员会 包括：拉斯·赫普尔博士、克里斯蒂安·莱温伯格和理查德·邓恩。他们对她充满了热情。 Sidhom是一名学生，并提供骨骼肌和成像方法方面的适当专业知识。西尔瓦那 已经完成了第一年的必修课程，因此委员会将与Sidhom女士合作，概述 一系列的教学课程，包括每年秋天教授的骨骼肌课程。该委员会也将提供帮助 指导Silvana进行体验式培训活动，使她掌握必要的技能，从而成为一名 独立的科学家，并完成国家卫生研究院的使命，推动不同群体的科学家的职业生涯。 该项目与母公司赠款的目标2直接相关： 特定目标2：测试时钟控制基因RBM20和/或TCAP在肌节中的作用 结构和肌肉功能。该目标将在BMal1缺陷的情况下使用TCAP和/或RBM20的AAV传递 以测试它们对肌节结构、肌原纤维蛋白组成和肌肉功能的贡献。 肌节结构元素将通过超分辨率显微镜和去卷积技术获得 用抗体定义2D和3D属性，包括肌节长度、粗丝中心性、M带 和Z线完整性。将对RBM20肌肉进行深度RNA测序，以确定剪接 改变。功能结果将包括体外措施，以测试结构变化对 被动/刚性和主动/最大力和收缩速率属性。

Abstract: The purpose of this project is to provide research and career development training for Silvana Sidhom, a current PhD student at the University of Florida. This administrative supplement in response to PA- 21-071: “Research Supplements to Promote Diversity in Health-Related Research (Admin Supp)” will facilitate career development for the candidate and as well as contribute to the mission of the parent project – to define molecular targets downstream of the circadian clock that modulate muscle structure with implications for strength. The primary objectives of Ms. Sidhom’s proposed research will be to work on experiments related to Aim 2 of the parent grant. First, she will learn to use confocal imaging approaches with sarcomeric antibodies to define changes in the M-line and Z-line structural elements of the sarcomere from the control and muscle specific Bmal1 KO mice. Second, she will use AAV approaches to restore expression of selected sarcomeric genes, such as Tcap, to test if that is sufficient to restore structure and muscle force. Ms. Sidhom will work actively with Collin Douglas, another PhD student working on the parent grant as well as Dr. Christopher Wolff, a postdoctoral associate on this project. Dr. Esser, the PI of the R01, will be the primary mentor for Ms. Sidhom within the College of Medicine at the University of Florida. We have established her committee and this includes: Drs. Russ Hepple, Christiaan Leeuwenburgh and Richard Dunn. They are enthusiastic about Ms. Sidhom as a student and provide appropriate expertise in skeletal muscle and imaging approaches. Silvana has completed her first year of required courses so the committee will work with Ms. Sidhom to outlined a series of didactic courses, including the skeletal muscle course taught each fall. The committee will also help guide Silvana with experiential training activities that will equip her with the necessary skills to progress as an independent scientist and to fulfill the NIH mission to advance the careers of a diverse population of scientists. The project is directly related to Aim 2 of the parent grant: Specific Aim 2: To test the clock controlled genes, Rbm20 and/or Tcap, for their roles in sarcomere structure and muscle function. This aim will use AAV delivery of Tcap and/or Rbm20 in Bmal1 deficient muscle to test their contribution to sarcomere structure, myofibrillar protein composition and muscle function. Sarcomere structural elements will be obtained by super-resolution microscopy with deconvolution techniques with antibodies to define 2D and 3D properties including sarcomere length, thick filament centrality, M-band and Z line integrity. Deep RNA-sequencing will be performed on the Rbm20 muscles to define splicing changes. Functional outcomes will include ex vivo measures to test the impact of structural changes on both passive/stiffness and active/max force and rate of contraction properties.