RGS14 integration of Gi/o and rap1/2 signaling pathways

超出提供的空间。神经递质和激素依赖于G蛋白在靶细胞上发挥作用。相反，生长因子酪氨酸激酶受体招募包括ras样GTP酶在内的一系列连接的信号蛋白来调节细胞的生长和分化。最近令人信服的证据表明，其他新的信号蛋白，包括G_1的某些调节因子。蛋白信号转导(RGS蛋白)在多个水平上整合了G蛋白和ras超家族调控通路。RGS蛋白是一种高度多样化的多功能蛋白，它直接与G蛋白结合来调节其功能。RGSI4包含我们和其他人已经证明与活性GCD/o-GTP(RGS结构域)、非活性GCD/o-GDP(GoLoco结构域)以及ras样GTP酶RAP1和Rap2(RID结构域)相互作用的结构域(RGS、GoLoco和RID结构域)。宿主细胞中RAPL/2通路的激活可刺激MAPK/Erk激酶信号转导和神经元分化。RGS14作为GAI和GAO信号的双重调节因子的作用尚不清楚。此外，RGS14、GCD/o和RAPL/2如何相互作用来调节其他功能，以及哪些细胞机制参与调节这些相互作用和相关的细胞反应尚不清楚。我们的初步研究表明，RGS14被参与G蛋白和RAP信号转导的激酶磷酸化。我们的假设是，RGS14是G蛋白(Ged/o)和RAP导向的信号通路的整合体，刺激启动的磷酸化调节RGS14与G_1和RAP的相互作用、其亚细胞定位及其信号功能。利用现代分子、细胞和生物化学手段，具体目标是：1.确定RGS I4作为受体和GOD和G(_O)信号在膜上的双功能调节器的作用。2.明确调控宿主B35细胞RGS 14膜募集和附着、亚细胞定位以及与mChoR、Gczi/o和RAPL/2结合的细胞和生化机制。3.明确RGS14在细胞中的磷酸化调控机制，以及RGS14在体外与mChoR/Gi/o和RAPT/2相互作用中的作用，以及B35细胞中相关的信号通路。4.确定在B35细胞中过表达RGSI4或消除天然RGS14对mChoR/Gi/o和RAPL/2导向的信号事件和细胞反应的影响。影响：这些研究将确定以前未知的RGS蛋白的细胞信号作用，并提供关于G蛋白信号通路和RAP/MAPKinase信号通路之间串扰的新机制的重要信息。这些研究的结果将澄清我们对G蛋白/RGS蛋白信号的复杂性的理解，并确定潜在的治疗干预的新靶点。PERFORMANCESITE(========================================Section End===========================================

EXCEED THE SPACE PROVIDED. Neurotransmitters and hormones rely upon G proteins to exert their effects on target cells. In contrast, growth factor tyrosine kinase receptors recruit a cascade of linked signaling proteins including ras-like GTPases to regulate cell growth and differentiation. Recent compelling evidence indicates that additional novel signaling proteins, including certain regulators of G_. protein signaling (RGS proteins) integrate G protein and ras superfamily-directed pathways at many levels. RGS proteins are highly diverse, multifunctional proteins that bind directly to G proteins to regulate their functions. RGSI4 contains domains (RGS, GoLoco and RID domains) that we and others have shown to interact with active Gcd/o-GTP (RGS domain), inactive Gcd/o-GDP (GoLoco domain) and with the ras-like GTPases rapl and rap2 (RID domain). Activation of rapl/2 pathways in host cells stimulates MAPK/Erk kinase signaling and neuronal differentiation. Roles for RGS14 as a dual regulator of Gai and Gao signaling are not known. Furthermore, how RGS14, Gcd/o and rapl/2 interact to regulate one anothers functions, and what cellular mechanisms are involved in regulating these interactions and linked cellular responses is not known. Our preliminary studies indicate that RGS14 is phosphorylated by kinases involved in G protein and rap signaling. Our hypothesis is that RGS14 is an integrator of G protein (Ged/o) and rap-directed signaling pathways, and that stimulus-initiated phosphorylation regulates RGS 14 interactions with G_ and rap, its subeellular localization, and its signaling functions. Using modern molecular, cellular and biochemical approaches, the specific aims will be to: 1. Determine roles for RGS I4 as a bifunctional regulator of receptor and God and G(_o signaling in membranes. 2. Define cellular and biochemical mechanisms that regulate RGS 14 membrane recruitment and attachment, subcellular localization, and a-Ssociati0n with mChoR, Gczi/o, and rapl/2 in host B35 cells. 3. Define cellular mechanisms for regulating RGS14 phosphorylation in cells, and roles for phosphorylation on RGS14 interactions with mChoR/Gi/o and rapt/2 in vitro, and linked signaling pathways in B35 cells. 4. Determine the effects of overexpressing RGSI4 or eliminating native RGS14 in B35 cells on mChoR/Gi/o and rapl/2- directed signaling events and cellular responses. Impact: These studies will identify previously unrecognized cell signaling roles for RGS proteins, and provide important information about novel mechanisms for cross-talk between G protein signaling pathways and rap/MAPkinase signaling pathways. Findings from these studies will clarify our understanding of the complexity of G protein/RGS protein signaling, and identify potential new targets for therapeutic intervention. PERFORMANCESITE( ========================================Section End===========================================