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===========================================

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Pharmacology A Study Section (PHRA)
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Stewart, Randall R
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Emory University
Schools of Medicine
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Gerber, Kyle J; Squires, Katherine E; Hepler, John R (2018) 14-3-3? binds regulator of G protein signaling 14 (RGS14) at distinct sites to inhibit the RGS14:G?i-AlF4- signaling complex and RGS14 nuclear localization. J Biol Chem 293:14616-14631
Squires, Katherine E; Gerber, Kyle J; Pare, Jean-Francois et al. (2018) Regulator of G protein signaling 14 (RGS14) is expressed pre- and postsynaptically in neurons of hippocampus, basal ganglia, and amygdala of monkey and human brain. Brain Struct Funct 223:233-253
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Zou, Juan; Salarian, Mani; Chen, Yanyi et al. (2017) Direct visualization of interaction between calmodulin and connexin45. Biochem J 474:4035-4051
Branch, Mary Rose; Hepler, John R (2017) Endogenous RGS14 is a cytoplasmic-nuclear shuttling protein that localizes to juxtanuclear membranes and chromatin-rich regions of the nucleus. PLoS One 12:e0184497
Brown, Nicole E; Lambert, Nevin A; Hepler, John R (2016) RGS14 regulates the lifetime of G?-GTP signaling but does not prolong G?? signaling following receptor activation in live cells. Pharmacol Res Perspect 4:e00249
Gerber, Kyle J; Squires, Katherine E; Hepler, John R (2016) Roles for Regulator of G Protein Signaling Proteins in Synaptic Signaling and Plasticity. Mol Pharmacol 89:273-86
Brown, Nicole E; Goswami, Devrishi; Branch, Mary Rose et al. (2015) Integration of G protein ? (G?) signaling by the regulator of G protein signaling 14 (RGS14). J Biol Chem 290:9037-49
Brown, Nicole E; Blumer, Joe B; Hepler, John R (2015) Bioluminescence resonance energy transfer to detect protein-protein interactions in live cells. Methods Mol Biol 1278:457-65

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