Signaling Pathways Regulating Proteolysis in Innervated Muscle

蛋白水解过程的调节在多种生理适应、发育和多种病理状态中是重要的，包括骨骼肌的消耗和萎缩以及心肌的肥大或萎缩。肌肉蛋白水解是众所周知的促进禁食或慢性饥饿，废用或去神经，在败血症和癌症恶病质。相反，减少蛋白水解（除了增加合成）可能有助于心脏肥大。通过细胞内或细胞外信号转导来调节肌肉中蛋白水解过程的机制知之甚少。这个项目的目的是增加我们对这些过程的理解，使用最简单的遗传学上易于处理的模型系统，用于研究神经支配的肌肉，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报告”蛋白质水解，特别是在95体壁肌肉和8性别肌肉细胞。报告蛋白在喂养的动物中是完全稳定的，但在饥饿、去神经支配、Ras原癌基因同源物的突变激活或成纤维细胞生长因子（FGF）受体同源物的突变超活化的条件下降解。本研究将确定激活的Ras信号对肌肉蛋白降解的控制是否由Raf/MEK/MAP激酶通路介导，确定其如何受P13激酶通路活性的影响，并确定来自激活的FGF受体的相关信号是否通过Ras/Raf/MEK/MAPK通路传递。肌内钙和甘油二酯的作用和钙激活蛋白激酶的可能作用也将进行探讨。这些问题将通过上位性分析的突变株，其中一个或两个信号转导蛋白被灭活或过度激活，结合使用选择性抑制剂的信号转导的各个步骤。在这个简单的系统中分析这些信号机制将为研究在更复杂的生物体中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.