Signaling Pathways Regulating Proteolysis in Innervated Muscle

蛋白质水解过程的调节在多种生理适应、发育和各种病理状态中都很重要，包括骨骼肌的消耗和萎缩，以及心肌的肥大或萎缩。众所周知，禁食或慢性饥饿、废用或去神经支配、败血症和癌症恶病质均可促进肌肉蛋白水解。相反，减少蛋白质水解（除了增加合成）可能导致心脏肥厚。细胞内或细胞外信号被转导调节肌肉蛋白水解过程的机制尚不清楚。这个项目的目的是增加我们对这些过程的理解，使用最简单的遗传可处理的模型系统来研究神经支配肌肉，秀丽隐杆线虫。该方法使用转基因动物，其中8-半乳糖苷酶融合蛋白的水平“在95个体壁肌和8个性肌细胞中特异性地进行了蛋白质水解”。报告蛋白在饲养动物中是完全稳定的，但在饥饿、去神经控制、Ras原癌基因同源基因的突变激活或成纤维细胞生长因子（FGF）受体同源基因的突变过度激活的条件下会降解。本研究将确定激活的Ras信号对肌肉蛋白降解的控制是否通过Raf/MEK/MAP激酶通路介导，确定其如何受到P13激酶通路活性的影响，并确定激活的FGF受体的相关信号是否通过Ras/Raf/MEK/MAPK通路传递。肌内钙和二酰基甘油的作用以及钙活化蛋白激酶的可能作用也将被探讨。这些问题将通过突变菌株的上位性分析来解决，其中一个或两个信号转导蛋白失活或过度活化，结合信号转导中单个步骤的选择性抑制剂的使用。在这个简单的系统中分析这些信号机制将为在更复杂的生物体中研究控制肌肉蛋白水解的信号通路提供范例。

Regulation of proteolytic processes is important in many kinds of physiological adaptations, in development, and in diverse pathological states, including wasting and atrophy in skeletal muscle, and hypertrophy or atrophy in cardiac muscle. Muscle proteolysis is well-known to be promoted by fasting or chronic starvation, by disuse or denervation, in sepsis and in cancer cachexia. Conversely, reduced proteolysis (in addition to increased synthesis) may contribute to cardiac hypertrophy. The mechanisms by which intracellular or extracellular signals are transduced to regulate proteolytic processes in muscle are poorly understood. This project aims to increase our understanding of these processes using the simplest genetically tractable model system available for studying innervated muscle, the nematode Caenorhabditis elegans. The methodology uses transgenic animals in which the level of a 8-galactosidase fusion protein reports" on proteolysis specifically in 95 body-wall muscle and 8 sex-muscle cells. The reporter protein is completely stable in fed animals, but is degraded under conditions of starvation, denervation, mutational activation of the homolog of the Ras proto-oncogene, or mutational hyperactivation of a fibroblast growth factor (FGF) receptor homolog. This research will determine whether control of muscle protein degradation by activated-Ras signals is mediated by the Raf/MEK/MAP kinase pathway, determine how it is affected by activity of the P13 kinase pathway, and determine if the relevant signals from activated FGF receptor are transmitted by way of the Ras/Raf/MEK/MAPK pathway. The roles of intramuscular calcium and diacylglycerol and the possible roles of calcium-activated protein kinases will also be explored. These questions will be approached by epistasis analysis of mutant strains in which one or two signal-transduction proteins are inactivated or hyperactivated, combined with the use of selective inhibitors of individual steps in signal transduction. Analysis of these signaling mechanisms in this simple system will provide paradigms for investigating the signaling pathways that control muscle proteolysis in more complex organisms.