title: Integration of G protein α (Gα) signaling by the regulator of G protein signaling 14 (RGS14).
abstract: RGS14 contains distinct binding sites for both active (GTP-bound) and inactive (GDP-bound) forms of Gα subunits. The N-terminal regulator of G protein signaling (RGS) domain binds active Gαi/o-GTP, whereas the C-terminal G protein regulatory (GPR) motif binds inactive Gαi1/3-GDP. The molecular basis for how RGS14 binds different activation states of Gα proteins to integrate G protein signaling is unknown. Here we explored the intramolecular communication between the GPR motif and the RGS domain upon G protein binding and examined whether RGS14 can functionally interact with two distinct forms of Gα subunits simultaneously. Using complementary cellular and biochemical approaches, we demonstrate that RGS14 forms a stable complex with inactive Gαi1-GDP at the plasma membrane and that free cytosolic RGS14 is recruited to the plasma membrane by activated Gαo-AlF4(-). Bioluminescence resonance energy transfer studies showed that RGS14 adopts different conformations in live cells when bound to Gα in different activation states. Hydrogen/deuterium exchange mass spectrometry revealed that RGS14 is a very dynamic protein that undergoes allosteric conformational changes when inactive Gαi1-GDP binds the GPR motif. Pure RGS14 forms a ternary complex with Gαo-AlF4(-) and an AlF4(-)-insensitive mutant (G42R) of Gαi1-GDP, as observed by size exclusion chromatography and differential hydrogen/deuterium exchange. Finally, a preformed RGS14·Gαi1-GDP complex exhibits full capacity to stimulate the GTPase activity of Gαo-GTP, demonstrating that RGS14 can functionally engage two distinct forms of Gα subunits simultaneously. Based on these findings, we propose a working model for how RGS14 integrates multiple G protein signals in host CA2 hippocampal neurons to modulate synaptic plasticity.
authors: From the Departments of Pharmacology and.
authors: Nicole E
authors: NE
authors: Brown
authors: the Department of Molecular Therapeutics, The Scripps Research Institute, Jupiter, Florida 33458.
authors: Devrishi
authors: D
authors: Goswami
authors: From the Departments of Pharmacology and.
authors: Mary Rose
authors: MR
authors: Branch
authors: From the Departments of Pharmacology and.
authors: Suneela
authors: S
authors: Ramineni
authors: Biochemistry, Emory University School of Medicine, Atlanta, Georgia 30322 and.
authors: Eric A
authors: EA
authors: Ortlund
authors: the Department of Molecular Therapeutics, The Scripps Research Institute, Jupiter, Florida 33458.
authors: Patrick R
authors: PR
authors: Griffin
authors: From the Departments of Pharmacology and jhepler@emory.edu.
authors: John R
authors: JR
authors: Hepler
mesh_terms: D000818:Animals; D001483:Base Sequence; D056651:CA2 Region, Hippocampal / Q000166:cytology / Q000378:metabolism; D017931:DNA Primers; D044385:GTP-Binding Protein alpha Subunits / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D009474:Neurons / Q000378:metabolism; D020710:RGS Proteins / Q000378:metabolism*; D051381:Rats; D015398:Signal Transduction
chemical_list: D017931:DNA Primers; D044385:GTP-Binding Protein alpha Subunits; D020710:RGS Proteins; C491400:RGS14 protein, human
keywords: G Protein; G Protein Regulatory (GPR) Motif; GTPase; GTPase-activating Protein (GAP); GoLoco Motif; RGS14; Regulator of G Protein Signaling (RGS); Synaptic Plasticity