INTERACTING PARTNERS OF G PROTEIN GAMMA SUBUNITS

G 蛋白伽马亚基的相互作用伙伴

• 批准号: 8365902
• 负责人: Stephen Farber
• 金额: $ 1.28万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8365902
• 关键词: Address; Affinity; Apoptosis; Binding; Biology; Chimera organism; Dominant-Negative Mutation; Event; Funding; Fungal Genome; G-Protein-Coupled Receptors; GTP-Binding Protein gamma Subunits; GTP-Binding Proteins; Grant; Heterotrimeric GTP-Binding Proteins; Hormonal; In Vitro; Individual; Mediating; National Center for Research Resources; Pathway interactions; Phototransduction; Principal Investigator; Protein Isoforms; Research; Research Infrastructure; Resources; Role; Signal Pathway; Signal Transduction; Source; Structure of primordial sex cell; Tertiary Protein Structure; United States National Institutes of Health; Vertebrates; cell motility; cost; in vivo; neurotransmission; prenylation; receptor-mediated signaling; response; zebrafish genome

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Signaling through heterotrimeric G proteins is essential for a variety of cellular responses including neurotransmission, hormonal response, olfactory transduction, phototransduction, cell migration and apoptosis. In response to the multitude of G protein coupled receptors (GPCR) that control these diverse pathways, vertebrates have evolved multiple isoforms of the alpha, beta and gamma subunits that selectively associate to form heterotrimeric G proteins. The zebrafish genome has 26alpha, 9beta and 17gamma subunits, which would make a total of 3978 potential different heterotrimers, if all combinations were made. Although the signaling partners and binding affinities of individual G protein isoforms have been investigated in vitro, the endogenous function of many heterotrimers and the extent of their functional redundancy remain uncharacterized. To address the functional diversity of heterotrimers containing specific Ggamma subunits in vivo, we expressed dominant negative versions of the Ggamma subunits to disrupt the signaling necessary for a known GPCR-mediated event, primordial germ cell (PGC) migration. We show that the vast majority of prenylation-deficient Ggamma subunits can disrupt PGC migration by altering the subcellular localization of signaling components. This disruption manifests in an inability of PGCs to migrate directionally. We identified a distinct subset of wild type Ggamma subunits that have the ability to reverse this semi-dominant negative effect, suggesting that Ggamma subunits have distinct and overlapping signaling capacities in vivo. To understand the roles Ggamma protein domains have in contributing to differences in Ggamma signaling capacity, we constructed Ggamma chimeras. Analysis of these chimeras demonstrated that multiple regions and motifs within the central and c-terminal regions influence the ability of Ggamma to mediate the signaling pathways necessary for directional PGC migration. Our results also indicate that prenylation-deficient Ggamma subunits can be used to disrupt GPCR-mediated signaling events in vivo.

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 通过异源三聚体G蛋白的信号传导对于多种细胞反应是必不可少的，包括神经传递、激素反应、嗅觉转导、光转导、细胞迁移和凋亡。响应于控制这些不同途径的大量G蛋白偶联受体（GPCR），脊椎动物已经进化出α、β和γ亚基的多种同种型，其选择性地缔合以形成异源三聚体G蛋白。斑马鱼的基因组有26个α、9个β和17个γ亚基，如果所有的组合都被制作出来，那么总共有3978个潜在的不同异源三聚体。虽然已在体外研究了单个G蛋白亚型的信号传导伙伴和结合亲和力，但许多异源三聚体的内源性功能及其功能冗余的程度仍不清楚。为了解决体内含有特定Ggamma亚基的异源三聚体的功能多样性，我们表达了Ggamma亚基的显性负性版本，以破坏已知GPCR介导的事件原始生殖细胞（PGC）迁移所必需的信号传导。我们发现，绝大多数的异戊烯基化缺陷Ggamma亚基可以破坏PGC迁移通过改变信号成分的亚细胞定位。这种破坏表现在PGCs不能定向迁移。我们鉴定了野生型Ggamma亚基的一个独特子集，其具有逆转这种半显性负效应的能力，表明Ggamma亚基在体内具有独特和重叠的信号传导能力。为了理解Ggamma蛋白结构域在Ggamma信号传导能力差异中的作用，我们构建了Ggamma嵌合体。对这些嵌合体的分析表明，中央和C-末端区域内的多个区域和基序影响G γ介导定向PGC迁移所需的信号传导途径的能力。我们的研究结果还表明，异戊烯基化缺陷的Ggamma亚基可用于破坏体内GPCR介导的信号转导事件。