title: Selective uncoupling of G alpha 12 from Rho-mediated signaling.
abstract: The heterotrimeric G protein G(12) has been implicated in such cellular regulatory processes as cytoskeletal rearrangement, cell-cell adhesion, and oncogenic transformation. Although the activated alpha-subunit of G(12) has been shown to interact directly with a number of protein effectors, the roles of many of these protein-protein interactions in G(12)-mediated cell physiology are poorly understood. To begin dissecting the specific cellular pathways engaged upon G(12) activation, we produced a series of substitution mutants in the regions of Galpha(12) predicted to play a role in effector binding. Here we report the identification and characterization of an altered form of Galpha(12) that is functionally uncoupled from signaling through the monomeric G protein Rho, a protein known to propagate several Galpha(12)-mediated signals. This mutant of Galpha(12) fails to bind the Rho-specific guanine nucleotide exchange factors p115RhoGEF and LARG (leukemia-associated RhoGEF), fails to stimulate Rho-dependent transcriptional activation, and fails to trigger activation of RhoA and the Rho-mediated cellular responses of cell rounding and c-jun N-terminal kinase activation. Importantly, this mutant of Galpha(12) retains coupling to the effector protein E-cadherin, as evidenced by its ability both to bind E-cadherin in vitro and to disrupt E-cadherin-mediated cell-cell adhesion. Furthermore, this mutant retains the ability to trigger beta-catenin release from the cytoplasmic domain of cadherin. This identification of a variant of Galpha(12) that is selectively uncoupled from one signaling pathway while retaining signaling capacity through a separate pathway will facilitate investigations into the mechanisms through which G(12) proteins mediate diverse biological responses.
authors.affiliation: Department of Pharmacology, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: Thomas E
authors.initials: TE
authors.lastname: Meigs
authors.forename: Juhi
authors.initials: J
authors.lastname: Juneja
authors.forename: C Todd
authors.initials: CT
authors.lastname: DeMarco
authors.forename: Laura N
authors.initials: LN
authors.lastname: Stemmle
authors.forename: Daniel D
authors.initials: DD
authors.lastname: Kaplan
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Casey
mesh_terms: D015820:Cadherins / Q000502:physiology; D002460:Cell Line; D044365:GTP-Binding Protein alpha Subunits, G12-G13 / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000502:physiology*; D006801:Humans; D011485:Protein Binding / Q000502:physiology; D064067:Rho Guanine Nucleotide Exchange Factors; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D015820:Cadherins; D020662:Guanine Nucleotide Exchange Factors; D064067:Rho Guanine Nucleotide Exchange Factors; D044365:GTP-Binding Protein alpha Subunits, G12-G13