Signaling Pathways Regulating Proteolysis in Innervated Muscle

这个项目的长期目标是了解调节肌肉中蛋白质降解的外部和肌肉内信号和信号转导网络。本实验室以前的研究已经发现并表征了蛋白酶体介导的由饥饿或功能性去神经支配引起的降解，以及成纤维细胞生长因子(成纤维细胞生长因子)通过Ras-MAP激酶级联信号诱导的非蛋白酶体降解，并显示了胰岛素样生长因子受体信号在对抗成纤维细胞生长因子促进的降解中的作用。这些研究表明，肌肉中的蛋白质降解不仅响应多种外部信号和自分泌信号，并由多种蛋白质分解机制介导，而且蛋白质分解代谢的状态可能取决于相反信号之间的平衡，而不是单一信号的强度。本研究旨在阐明秀丽线虫肌肉中转基因报告蛋白的降解受转化生长因子β受体(TGF-β)负调控，并受钙钙调素依赖蛋白激酶(CaMKII)过度信号的促进。使用的方法主要是遗传学(特别是抑制和上位性分析)、生物化学(报告蛋白的测量和信号的分子相关性)和分子生物学。潜在的新的信号通路将被阐明，转化生长因子-β受体和CaM激酶下游的效应器将被表征，这些通路与细胞中其他通路的关系将被探索。这个项目的更广泛的影响，在短期到中期，将提供一个更敏锐的理解复杂的信号和信号转导网络，至少在两种不同的细胞类型(神经元和肌肉)和可能的第三种细胞(皮下组织)中运作，以调节目标组织(肌肉)中的蛋白质稳定性。这也将产生重大的教育影响：本科生研究人员将成为项目团队不可或缺的一部分。高级科学人员将作出巨大努力，将线虫生物学和神经肌肉生物学的各个方面纳入他们教授的本科课程(遗传学、生物学入门)和其他本科课程(发育生物学、细胞生物学)。将线虫作为胰岛素信号和脂肪代谢的模型的模块被纳入高中教师的研讨会，并为K-12学校的访问提供了活的线虫。从长远来看，这个项目对临床上重要的肌肉健康问题的影响可能会很大，因为肌肉蛋白质的分解代谢对正常的适应性生理学和各种病理状态都是重要的。

The long-range objective of this project is to understand the external and intramuscular signals and signal-transduction networks that regulate protein degradation in muscle. Previous studies in this laboratory have identified and characterized the proteasome-mediated degradation induced by starvation or functional denervation and the non-proteasomal degradation induced by Fibroblast Growth Factor (FGF) signaling via a Ras-MAP kinase cascade, and shown a role of Insulin-like Growth Factor receptor signaling in opposing FGF-promoted degradation. These studies made it clear not only that protein degradation in muscle responds to multiple external and autocrine signals and is mediated by multiple proteolytic mechanisms, but also that the state of protein catabolism may be determined by the balance between opposing signals rather than by the intensity of a single signal. This proposal focuses on elucidating the observations that degradation of transgene-coded reporter proteins in muscles of Caenorhabditis elegans is negatively regulated by a Transforming Growth Factor Beta (TGF-beta) receptor, and is promoted by excess signal from a calcium-calmodulin dependent protein kinase (CaMKII). The methodologies used are primarily those of genetics (particularly suppression and epistasis analysis), biochemistry (measurement of reporter proteins and molecular correlates of signaling) and molecular biology. Potentially novel signaling pathways will be elucidated, effectors downstream of the TGF-beta receptor and CaM kinase will be characterized, and the relation of these pathways to others in the cell will be explored. The broader impact of this project, in the short-to-medium term, will be to provide a keener understanding of how complex networks of signals and signal-transduction pathways, operating in at least two distinct cell types (neurons and muscles) and possibly a third (hypodermis), are coordinated to regulate protein stability in the target tissue (muscle). There will also be significant educational impacts: Undergraduate researchers will be an integral part of the project team. The senior scientific personnel will make strong efforts to incorporate aspects of C. elegans biology and neuromuscular biology in general into undergraduate courses they teach (Genetics, Introductory Biology) and into other undergraduate courses (Developmental Biology, Cell Biology). Modules using C. elegans as a model for insulin signaling and lipid metabolism are incorporated into workshops for high-school teachers and live C. elegans are provided for K-12 school visits. In the longer term, the impact of this project on clinically important issues of muscle health could become substantial, since the catabolism of muscle proteins is important both for normal adaptive physiology and for a variety of pathological states.