title: Receptor-coupling properties of the invertebrate visual guanine nucleotide binding protein iGqalpha.
abstract: Invertebrate visual iG(q)alpha is homologous to mammalian mG(q)alpha in two of the three domains important for G protein interaction with receptors; the C-terminus and the linker regions that connect the helical and ras-like domains of the alpha subunit. The third receptor-interacting domain, the N-terminus, contains a six amino acid extension MTLESI in mG(q)alpha that is not present in iG(q)alpha. In co-expression studies we assessed the promiscuity and efficacy of receptor coupling to phospholipase C (PLC) by iG(q)alpha, a non-palmitoylated mutant iG(q)alpha(C3,4A), mG(q)alpha and G(15)alpha. The invertebrate G proteins and mG(q)alpha only coupled to G(q)-coupled receptors (m1 muscarinic acetylcholine receptor (mChR1), alpha(1A)-adrenergic receptor (alpha1-AR)) and not to the G(i/s)-coupled receptors (CCR1 receptor, beta2-adrenergic receptor or dopamine D1 receptor) while G(15)alpha coupled to all receptors. iG(q)alpha and iG(q)alpha(C3,4A) both had double the efficacy for PLC activation compared to the mammalian G proteins when co-expressed with mChR1 and alpha1-AR. The increased efficacy of iG(q)alpha compared to mG(q)alpha was also seen downstream of PLC with carbachol stimulation of the mitogen-activated protein kinase, ERK1/2. Addition of the MTLESI extension onto the N-terminus of iG(q)alpha decreased its efficacy by 35% whereas deletion of this sequence from mG(q)alpha increased its efficacy by 60% in the PLC and ERK1/2 assays. iG(q)alpha, iG(q)alpha(C3,4A) and mG(q)alpha all displayed similar receptor-independent AlF(4)(-)activation of PLC and guanosine triphosphate hydrolysis (GTPase) activity. iG(q)alpha, and iG(q)alpha(C3,4A) both had increased receptor-activated guanosine 5'-[gamma-[(35)S]thio]triphosphate ([(35)S]GTPgammaS) binding when compared to mG(q)alpha when co-expressed with the mChR1. These results demonstrate that G(q) protein efficacy is at least partially determined by the presence of the amino-terminal MTLESI extension. Comparison of [(35)S]GTPgammaS binding rates helps explain the increased efficacy of the invertebrate G proteins.
authors.affiliation: Department of Pharmacology, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Lynle
authors.initials: L
authors.lastname: Go
authors.forename: Jane
authors.initials: J
authors.lastname: Mitchell
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D004789:Enzyme Activation / Q000187:drug effects; D005136:Eye Proteins / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000378:metabolism; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000737:chemistry / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006801:Humans; D006868:Hydrolysis / Q000187:drug effects; D049834:Loligo / Q000378:metabolism*; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D043643:Receptor, Muscarinic M1 / Q000378:metabolism; D018340:Receptors, Adrenergic, alpha-1 / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry / Q000378:metabolism*; D013329:Structure-Activity Relationship; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D005136:Eye Proteins; D050505:Mutant Proteins; D043643:Receptor, Muscarinic M1; D018340:Receptors, Adrenergic, alpha-1; D043562:Receptors, G-Protein-Coupled; D006160:Guanosine Triphosphate; D002217:Carbachol; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D010738:Type C Phospholipases; D020558:GTP Phosphohydrolases; D043802:GTP-Binding Protein alpha Subunits, Gq-G11