RGS-insensitive Gα subunits: probes of Gα subtype-selective signaling and physiological functions of RGS proteins.
ABSTRACT The Regulator of G protein Signaling (RGS) proteins were identified as a family in 1996 and humans have more than 30 such proteins. Their best known function is to suppress G Protein-Coupled Receptors (GPCR) signaling by increasing the rate of Gα turnoff through stimulation of GTPase activity (i.e., GTPase acceleration protein or GAP activity). The GAP activity of RGS proteins on the Gαi and Gαq family of G proteins can terminate signals initiated by both α and βγ subunits. RGS proteins also serve as scaffolds, assembling signal-regulating modules. Understanding the physiological roles of RGS proteins is of great importance, as GPCRs are major targets for drug development. The traditional method of using RGS knockout mice has provided some information about the role of RGS proteins but in many cases effects are modest, perhaps because of redundancy in RGS protein function. As an alternative approach, we have utilized a glycine-to-serine mutation in the switch 1 region of Gα subunits that prevents RGS binding. The mutation has no known effects on Gα binding to receptor, Gβγ, or effectors. Alterations in function resulting from the G>S mutation imply a role for both the specific mutated Gα subunit and its regulation by RGS protein activity. Mutant rodents expressing these G>S mutant Gα subunits have strong phenotypes and provide important information about specific physiological functions of Gαi2 and Gαo and their control by RGS. The conceptual framework behind this approach and a summary of recent results is presented in this chapter.
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ABSTRACT: Regulators of G protein signaling (RGS) proteins are emerging as potentially important drug targets. The mammalian RGS protein family has more than 20 members and they share a common ∼120-residue RGS homology domain or "RGS box." RGS proteins regulate signaling via G protein-coupled receptors by accelerating GTPase activity at active α subunits of G proteins of the G(q) and G(i/o) families. Most studies searching for modulators of RGS protein function have been focused on inhibiting the GTPase accelerating protein activity. However, many RGS proteins contain additional domains that serve other functions, such as interactions with proteins or subcellular targeting. Here, we discuss a rationale for therapeutic targeting of RGS proteins by regulation of expression or allosteric modulation to permit either increases or decreases in RGS function. Several RGS proteins have reduced expression or function in pathophysiological states, so strategies to increase RGS function would be useful. Because several RGS proteins are rapidly degraded by the N-end rule pathway, finding ways to stabilize them may prove to be an effective way to enhance RGS protein function.Molecular pharmacology 10/2010; 78(4):550-7. DOI:10.1124/mol.110.065219 · 4.12 Impact Factor
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ABSTRACT: Heterotrimeric G-proteins are molecular switches integral to a panoply of different physiological responses that many organisms make to environmental cues. The switch from inactive to active Gαβγ heterotrimer relies on nucleotide cycling by the Gα subunit: exchange of GTP for GDP activates Gα, whereas its intrinsic enzymatic activity catalyzes GTP hydrolysis to GDP and inorganic phosphate, thereby reverting Gα to its inactive state. In several genetic studies of filamentous fungi, such as the rice blast fungus Magnaporthe oryzae, a G42R mutation in the phosphate-binding loop of Gα subunits is assumed to be GTPase-deficient and thus constitutively active. Here, we demonstrate that Gα(G42R) mutants are not GTPase deficient, but rather incapable of achieving the activated conformation. Two crystal structure models suggest that Arg-42 prevents a typical switch region conformational change upon Gα(i1)(G42R) binding to GDP·AlF(4)(-) or GTP, but rotameric flexibility at this locus allows for unperturbed GTP hydrolysis. Gα(G42R) mutants do not engage the active state-selective peptide KB-1753 nor RGS domains with high affinity, but instead favor interaction with Gβγ and GoLoco motifs in any nucleotide state. The corresponding Gα(q)(G48R) mutant is not constitutively active in cells and responds poorly to aluminum tetrafluoride activation. Comparative analyses of M. oryzae strains harboring either G42R or GTPase-deficient Q/L mutations in the Gα subunits MagA or MagB illustrate functional differences in environmental cue processing and intracellular signaling outcomes between these two Gα mutants, thus demonstrating the in vivo functional divergence of G42R and activating G-protein mutants.PLoS Pathogens 02/2012; 8(2):e1002553. DOI:10.1371/journal.ppat.1002553 · 8.06 Impact Factor
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