Stress-induced myosin folding and assembly mechanisms

应激诱导的肌球蛋白折叠和组装机制

• 批准号: 401331821
• 负责人: Professor Dr. Thorsten Hoppe
• 金额: --
• 依托单位: Institut für Genetik
• 依托单位国家: 德国
• 项目类别: Research Units
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/401331821?language=en
• 关键词: Stress; induced; myosin; folding; assembly

Muscle function and maintenance is governed by the spatiotemporal organization of structural and motor proteins into contractile sarcomeres. Given the constant use and mechanical stress exposure of the muscle tissue, the sarcomeric integrity is continuously protected by a complex network of protein folding and degradation pathways. An important regulator of sarcomere formation and muscle function is the HSP90 co-chaperone UNC-45. According to its central role in myosin assembly, UNC-45 protein amount and localization are tightly coordinated with mechanical stress conditions or induced damage to the myofiber. We are particularly interested in the myosin-directed UNC-45 chaperone network, orchestrating myofibrillogenesis from Caenorhabditis elegans to man. Besides elucidating the disease relevance of the human orthologue UNC-45B, we successfully developed powerful experimental approaches during the first funding period for the in-depth investigation of muscle-specific quality control pathways. In order to define myosin-directed stress response mechanisms, we have established optogenetic methods, transgenic reporter assays, and electrical pulse stimulation both in worms and murine myotubes in combination with optimized pull-down and mass spectrometry protocols. Moreover, we identified a so far undiscovered muscle-specific waste management pathway that is regulated by UNC-45. The central objective of the proposed research is to understand how protein folding and degradation networks are coordinated with the dynamics of muscle organization and repair in the context of mechanical stress conditions. The suggested project will address myosin-directed stress response programs and systematically analyze: the role of UNC-45 in protein synthesis, protein degradation, and exopher formation (Aim 1), mechanical stress-induced UNC-45 regulation and myosin-directed quality control (Aim 2), and the conserved regulation of UNC-45 function (Aim 3). To this end, optogenetic induction of mechanical stress, proximity labeling technologies, CRISPR/Cas9 gene editing, automated locomotor activity measurement, and large-scale genetic screenings will be performed. Given our recent discoveries of myopathy-related UNC-45 mutations and a novel muscle-specific waste management system, combined with state-of-the-art technologies established during the first funding period, we are ideally positioned to define mechanical stress response mechanisms that protect muscle function during acute physical exercise or continued use.

肌肉功能和维持由结构蛋白和运动蛋白在收缩肌节中的时空组织控制。鉴于肌肉组织的持续使用和机械应力暴露，肌节的完整性受到蛋白质折叠和降解途径的复杂网络的持续保护。肌节形成和肌肉功能的重要调节因子是 HSP90 共伴侣 UNC-45。根据其在肌球蛋白组装中的核心作用，UNC-45 蛋白的数量和定位与机械应力条件或诱发的肌纤维损伤紧密协调。我们对肌球蛋白引导的 UNC-45 伴侣网络特别感兴趣，该网络协调从秀丽隐杆线虫到人类的肌原纤维发生。除了阐明人类直系同源物 UNC-45B 的疾病相关性外，我们还在第一个资助期内成功开发了强大的实验方法，用于深入研究肌肉特异性质量控制途径。为了定义肌球蛋白导向的应激反应机制，我们在线虫和小鼠肌管中建立了光遗传学方法、转基因报告分析和电脉冲刺激，并结合优化的下拉和质谱方案。此外，我们还发现了迄今为止尚未发现的受 UNC-45 监管的肌肉特异性废物管理途径。拟议研究的中心目标是了解蛋白质折叠和降解网络如何在机械应力条件下与肌肉组织和修复的动力学相协调。建议的项目将解决肌球蛋白导向的应激反应计划并系统分析：UNC-45 在蛋白质合成、蛋白质降解和外层形成中的作用（目标 1）、机械应力诱导的 UNC-45 调节和肌球蛋白导向的质量控制（目标 2）以及 UNC-45 功能的保守调节（目标 3）。为此，将进行机械应力的光遗传学诱导、邻近标记技术、CRISPR/Cas9基因编辑、自动运动活动测量和大规模遗传筛选。鉴于我们最近发现的与肌病相关的 UNC-45 突变和新型肌肉特异性废物管理系统，再加上第一个资助期间建立的最先进的技术，我们处于理想的位置，可以定义在急性体育锻炼或持续使用期间保护肌肉功能的机械应力反应机制。