Molecular mechanisms of ARF family GTPases

项目摘要/摘要 ARF调节GTP酶家族的成员在细胞信号转导中发挥节点的作用 协调重要的细胞过程，包括膜运输、能量代谢、 纤毛发生和细胞骨架。我首先研究了ARF和后来的ARF样蛋白(ARL) 30多年来，使用生化、细胞和分子生物学、遗传学和 并建议继续进行这些研究，重点放在它们的行动上 连接到纤毛和中心体，以允许对五个子集进行更详细的机制研究 这个家族中有30个GTP酶。我们将使用基因组编辑来生成一组基因敲除 通过研究中的蛋白质破译信号的细胞系。我们将使用经典蛋白质 生物化学来提纯和发现这些途径中的新成分。我们都会学习 这五种GTP酶在细胞中的功能，并与它们的体外生化研究相联系 酶学。我们将探索细胞信号的串扰或更高级别排序的可能性， 还开发了针对非典型GTP酶的新的、单一的作用，作为确定共享的手段 或家庭内部独特的机制。对这些系统的更好理解将揭示 对细胞生物学基本方面的新见解，以及为 干预以改变人类疾病的进程；包括但不限于癌症、心脏 疾病、神经退行性变、纤毛病变、视网膜退行性变和耳聋。

Project Summary/Abstract Members of the ARF family of regulatory GTPases function as nodes in cell signaling to coordinate essential cell processes; including membrane traffic, energy metabolism, ciliogenesis, and the cytoskeleton. I have studied first ARF and later ARF-like (ARL) proteins for over 30 years, using a combination of biochemical, cell and molecular biological, genetic, and phylogenetic approaches and propose to continue these studies with a focus on their actions linked to cilia and centrosomes to allow more detailed mechanistic studies of a subset of five of the 30 GTPases in this family. We will use genome editing to generate a collection of knockout cell lines to decipher signaling by the proteins under study. And we will use classical protein biochemistry to purify and discover novel components in these pathways. We will both study functions of each of these five GTPases in cells, and link to in vitro biochemical studies of their enzymology. We will explore the potential for cross-talk or higher level ordering of cell signaling, and also develop novel, single actions for atypical GTPases as a means of determining shared or unique mechanisms within the family. A better understanding of these systems will reveal novel insights into fundamental aspects of cell biology as well as providing potential targets for intervention to alter the course of human diseases; including but not limited to cancer, heart disease, neurodegeneration, ciliopathies, retinal degeneration, and deafness.