Unraveling the regulation of MuRF1 activity, a central hub in a dynamic machinery thatpromotes skeletal muscle atrophy.

揭示 MuRF1 活性的调节，MuRF1 是促进骨骼肌萎缩的动态机制的中心枢纽。

• 批准号: 495189339
• 负责人: Professor Dr. Jens Fielitz
• 金额: --
• 依托单位: Forschungsgruppe Intrazelluläre Proteolyse
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/495189339?language=en
• 关键词: Unraveling; regulation; MuRF1; activity; central

Loss of skeletal muscle mass in response to reduced physical strain, malnutrition, or other factors is a health threat for diseased or elderly people. Therefore, decreasing the rate of muscle wasting is considered to alleviate the morbidity and mortality of patients. During muscle wasting, also termed muscle atrophy, a highly ordered functional structure within myofibers called the sarcomere is degraded and this process involves the ubiquitin proteasome system (UPS). Indeed, two bona fide components of the UPS, the “Muscle RING Finger 1” (MuRF1; encoded by Trim63) and Atrogin-1 (encoded by Fbxo32) have been identified, which both are exclusively expressed in muscle cells and which both were shown to be required for the atrophic process. Still, relatively few is known about the cellular function of MuRF1 and Atrogin-1 and their molecular activities during muscle wasting. In a recent work, we identified MuRF1 as a component of a cullin-type ubiquitin ligase. These multi-enzyme protein complexes are built around a cullin scaffold unit, which binds different combinations of substrate recruitment factors and a RING protein. Due to their modular character, cullin-type ubiquitin ligases mediate the ubiquitination of highly diverse client proteins and adapt their activity to changing requirements by replacing individual substrate binding subunits. Various high-throughput screens led to the identification of putative MuRF1 substrates such as myosin heavy chain (MyHC), myosin light chain 2, titin, muscle-type creatine kinase, and glucocorticoid modulatory element binding protein-1, but it is unclear, whether ubiquitination of these proteins involves MuRF1. This research plan aims to better characterize the function of MuRF1 in skeletal muscle atrophy. We will analyze already available high-throughput transcriptomics and proteomics data to study the regulation of MuRF1 and known partner proteins during atrophy and identify novel interacting factors as well as substrate proteins. We then want to employ biochemical methods to investigate the function of these proteins in cell culture. The activity and regulation of selected candidates will then be studied in an animal system for muscle atrophy. Finally, we plan to reconstitute MuRF1 containing protein complexes with purified components and monitor their ubiquitination activity in vitro. The biological significance of our findings will then be verified using patient material. This work should provide deeper insights into the molecular activities of MuRF1 and its partner proteins during skeletal muscle atrophy and possibly unravel novel entry points for clinical intervention.

由于身体压力减轻、营养不良或其他因素而导致的骨骼肌质量损失对患病或老年人的健康构成威胁。因此，降低肌肉萎缩率被认为可以减轻患者的发病率和死亡率。在肌肉萎缩（也称为肌肉萎缩）期间，肌纤维内称为肌节的高度有序的功能结构被降解，该过程涉及泛素蛋白酶体系统（UPS）。事实上，已经鉴定出 UPS 的两个真正成分，即“肌肉环指 1”（MuRF1；由 Trim63 编码）和 Atrogin-1（由 Fbxo32 编码），它们都专门在肌肉细胞中表达，并且都被证明是萎缩过程所必需的。然而，对于 MuRF1 和 Atrogin-1 的细胞功能及其在肌肉萎缩过程中的分子活性知之甚少。在最近的一项工作中，我们确定 MuRF1 是 cullin 型泛素连接酶的一个组成部分。这些多酶蛋白复合物围绕 cullin 支架单元构建，该单元结合底物招募因子和 RING 蛋白的不同组合。由于其模块化特征，cullin 型泛素连接酶介导高度多样化的客户蛋白的泛素化，并通过替换单个底物结合亚基来调整其活性以适应不断变化的需求。各种高通量筛选鉴定了假定的 MuRF1 底物，例如肌球蛋白重链 (MyHC)、肌球蛋白轻链 2、肌联蛋白、肌肉型肌酸激酶和糖皮质激素调节元件结合蛋白 1，但尚不清楚这些蛋白质的泛素化是否涉及 MuRF1。该研究计划旨在更好地表征 MuRF1 在骨骼肌萎缩中的功能。我们将分析现有的高通量转录组学和蛋白质组学数据，以研究 MuRF1 和已知伙伴蛋白在萎缩过程中的调节，并识别新的相互作用因子以及底物蛋白。然后我们想采用生化方法来研究这些蛋白质在细胞培养物中的功能。然后将在肌肉萎缩的动物系统中研究选定候选者的活动和调节。最后，我们计划用纯化的成分重建含有 MuRF1 的蛋白质复合物，并监测其体外泛素化活性。我们的研究结果的生物学意义将使用患者材料进行验证。这项工作应该可以更深入地了解 MuRF1 及其伙伴蛋白在骨骼肌萎缩过程中的分子活性，并可能揭示临床干预的新切入点。