Activation of nematode G protein GOA-1 by the human muscarinic acetylcholine receptor M2 subtype. Functional coupling of G-protein-coupled receptor and G protein originated from evolutionarily distant animals.
Signal transduction mediated by heterotrimeric G proteins regulates a wide variety of physiological functions. We are interested in the manipulation of G-protein-mediating signal transduction using G-protein-coupled receptors, which are derived from evolutionarily distant organisms and recognize unique ligands. As a model, we tested the functionally coupling GOA-1, G alpha(i/o) ortholog in the nematode Caenorhabditis elegans, with the human muscarinic acetylcholine receptor M2 subtype (M2), which is one of the mammalian G alpha(i/o)-coupled receptors. GOA-1 and M2 were prepared as a fusion protein using a baculovirus expression system. The affinity of the fusion protein for GDP was decreased by addition of a muscarinic agonist, carbamylcholine and the guanosine 5'-[3-O-thio]triphosphate ([35S]GTPgammaS) binding was increased with an increase in the carbamylcholine concentrations in a dose-dependent manner. These effects evoked by carbamylcholine were completely abolished by a full antagonist, atropine. In addition, the affinity for carbamylcholine decreased under the presence of GTP as reported for M2-G alpha(i/o) coupling. These results indicate that the M2 activates GOA-1 as well as G alpha(i/o).
Minaba
Masaomi
M
Immune Defense Unit, National Institute of Agrobiological Sciences, Ibaraki, Japan.
Ichiyama
Susumu
S
Kojima
Katsura
K
Ozaki
Mamiko
M
Kato
Yusuke
Y
D000595:Amino Acid Sequence; D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics*; D029742:Caenorhabditis elegans Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D019143:Evolution, Molecular; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D019204:GTP-Binding Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D018721:Muscarinic Agonists / Q000494:pharmacology; D011485:Protein Binding / Q000235:genetics; D043585:Receptor, Muscarinic M2 / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment
D029742:Caenorhabditis elegans Proteins; C516620:GOA-1 protein, C elegans; D018721:Muscarinic Agonists; D043585:Receptor, Muscarinic M2; D011993:Recombinant Fusion Proteins; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins; D019206:GTP-Binding Protein alpha Subunits, Gi-Go