title: Reversible translocation of p115-RhoGEF by G(12/13)-coupled receptors.
abstract: G protein-coupled receptors (GPCRs) are important targets for medicinal agents. Four different G protein families, G(s), G(i), G(q), and G(12), engage in their linkage to activation of receptor-specific signal transduction pathways. G(12) proteins were more recently studied, and upon activation by GPCRs they mediate activation of RhoGTPase guanine nucleotide exchange factors (RhoGEFs), which in turn activate the small GTPase RhoA. RhoA is involved in many cellular and physiological aspects, and a dysfunction of the G(12/13)-Rho pathway can lead to hypertension, cardiovascular diseases, stroke, impaired wound healing and immune cell functions, cancer progression and metastasis, or asthma. In this study, regulator of G protein signaling (RGS) domain-containing RhoGEFs were tagged with enhanced green fluorescent protein (EGFP) to detect their subcellular localization and translocation upon receptor activation. Constitutively active Galpha(12) and Galpha(13) mutants induced redistribution of these RhoGEFs from the cytosol to the plasma membrane. Furthermore, a pronounced and rapid translocation of p115-RhoGEF from the cytosol to the plasma membrane was observed upon activation of several G(12/13)-coupled GPCRs in a cell type-independent fashion. Plasma membrane translocation of p115-RhoGEF stimulated by a GPCR agonist could be completely and rapidly reversed by subsequent application of an antagonist for the respective GPCR, that is, p115-RhoGEF relocated back to the cytosol. The translocation of RhoGEF by G(12/13)-linked GPCRs can be quantified and therefore used for pharmacological studies of the pathway, and to discover active compounds in a G(12/13)-related disease context.
authors.affiliation: Center for Proteomic Chemistry, Novartis Institutes for BioMedical Research Basel, Novartis Pharma AG, 4002 Basel, Switzerland.
authors.forename: Bruno H
authors.initials: BH
authors.lastname: Meyer
authors.forename: Felix
authors.initials: F
authors.lastname: Freuler
authors.forename: Danilo
authors.initials: D
authors.lastname: Guerini
authors.forename: Sandra
authors.initials: S
authors.lastname: Siehler
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D004285:Dogs; D044365:GTP-Binding Protein alpha Subunits, G12-G13 / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D021381:Protein Transport; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D049349:Receptors, Lysosphingolipid / Q000037:antagonists & inhibitors; D064067:Rho Guanine Nucleotide Exchange Factors; D013347:Subcellular Fractions
chemical_list: C576600:ARHGEF1 protein, human; D020662:Guanine Nucleotide Exchange Factors; D043562:Receptors, G-Protein-Coupled; D049349:Receptors, Lysosphingolipid; D064067:Rho Guanine Nucleotide Exchange Factors; D044365:GTP-Binding Protein alpha Subunits, G12-G13