Mechanisms of RGS Function in Vertebrate Development

RGS 在脊椎动物发育中的功能机制

• 批准号: 6830801
• 负责人: DIANE C SLUSARSKI
• 金额: $ 7.38万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-12-01 至 2005-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6830801
• 关键词: G protein; biological signal transduction; calcium flux; developmental genetics; fluorescent dye /probe; genetically modified animals; guanosinetriphosphatase activating protein; in situ hybridization; nonmammalian vertebrate embryology; protein structure function; proteomics; zebrafish

DESCRIPTION (provided by applicant): Heterotrimeric G proteins represent an extremely common and important component of intracellular signaling in organisms ranging from yeast to man, functioning to transduce signals from specific G protein coupled receptors to effector proteins. Regulators of G protein signaling (RGS) are GTPase-activating proteins (GAPs) for certain G-alpha subunits and, therefore, possess the potential to regulate the active lifetimes of both G-alpha and G-beta-gamma subunits in vivo. However, little is known about the in vivo functions of RGS proteins in vertebrates apart from their role in regulating phototransduction in the retina. Here the investigators report the first evidence for RGS function in zebrafish development. Misexpression of human RGS16 (hRGS16) in zebrafish embryos produced marked phenotypic defects in neural patterning and cell motility that were intriguingly similar to Wnt-induced defects. Additionally, loss of zebrafish RGS3s (zRGS3s) function results in somite patterning defects. This proposal is designed to perform a comprehensive analysis of the role of RGS proteins in vertebrate developmental processes utilizing the experimentally amenable zebrafish. RGS proteins are a large diverse family of multi-functional signaling proteins that have been divided into subgroups. Members of the R4 subgroup (including RGS1, 2, 3, 4, 5, 8, 13, 16 and 18) are proposed to act exclusively as negative regulators of G-protein function. In zebrafish, the investigators have identified expressed sequence tags encoding all nine members of the R4 subfamily of RGS proteins as well as G-alpha proteins. Insight into functional roles and potential functional differences among RGS proteins may be reflected by their cellular distribution. The investigators will characterize patterns of expression of all zRGS R4 family members and determine the developmental consequences of their loss-of-function by gene knockdown studies. The investigators will also test the hypothesis that the intrinsic GAP activity on G proteins may target Wnt/fz signaling. The Wnt/fz signal transduction pathway is a key developmental pathway, which the investigators have demonstrated requires participation of a PTX-sensitive G protein and has been implicated in several human cancers. Demonstrating an impact of RGS proteins on this pathway will represent a major advance in our understanding of the Wnt/fz signal transduction pathway. Some RGS proteins may use such a mechanism while others may not, however the proposed strategy offers the unprecedented opportunity to elucidate the developmental role of RGS proteins with both gain and loss of function analyses in the vertebrate embryo.

描述（由申请人提供）：异三聚体G蛋白代表了从酵母到人类的生物体中细胞内信号传导的极其常见和重要的成分，其功能是将信号从特定G蛋白偶联受体转导至效应蛋白。 G 蛋白信号传导 (RGS) 调节剂是某些 G-α 亚基的 GTP 酶激活蛋白 (GAP)，因此具有调节体内 G-α 和 G-β-γ 亚基活性寿命的潜力。 然而，除了 RGS 蛋白在调节视网膜光转导中的作用之外，人们对脊椎动物体内 RGS 蛋白的功能知之甚少。在这里，研究人员报告了 RGS 在斑马鱼发育中发挥作用的第一个证据。斑马鱼胚胎中人类 RGS16 (hRGS16) 的错误表达会在神经模式和细胞运动方面产生明显的表型缺陷，这些缺陷与 Wnt 诱导的缺陷非常相似。此外，斑马鱼 RGS3s (zRGS3s) 功能的丧失会导致体节图案缺陷。 该提案旨在利用经过实验验证的斑马鱼，对 RGS 蛋白在脊椎动物发育过程中的作用进行全面分析。 RGS 蛋白是一个大而多样化的多功能信号蛋白家族，已分为多个亚组。 R4 亚组的成员（包括 RGS1、2、3、4、5、8、13、16 和 18）被认为专门充当 G 蛋白功能的负调节因子。在斑马鱼中，研究人员已经鉴定出编码 RGS 蛋白 R4 亚家族所有九个成员以及 G-α 蛋白的表达序列标签。对 RGS 蛋白的功能作用和潜在功能差异的深入了解可以通过它们的细胞分布来反映。研究人员将表征所有 zRGS R4 家族成员的表达模式，并通过基因敲除研究确定其功能丧失的发育后果。研究人员还将测试 G 蛋白的内在 GAP 活性可能针对 Wnt/fz 信号传导的假设。 Wnt/fz 信号转导途径是一条关键的发育途径，研究人员已证明该途径需要 PTX 敏感 G 蛋白的参与，并且与多种人类癌症有关。证明 RGS 蛋白对该通路的影响将代表我们对 Wnt/fz 信号转导通路理解的重大进展。一些 RGS 蛋白可能使用这种机制，而另一些则可能不会，但是所提出的策略提供了前所未有的机会，通过脊椎动物胚胎中功能的获得和丧失分析来阐明 RGS 蛋白的发育作用。