Identification of G protein alpha-subunits in RINm5F cells and their selective interaction with galanin receptor.
Galanin, an inhibitor of insulin secretion in pancreatic beta-cells, exerts its multiple effects through mechanisms that are sensitive to pertussis toxin (PTX). G proteins have been characterized in RINm5F cells. By ADP ribosylation and immunoblotting, the alpha-subunits of Gi1, Gi2, Gi3, and two forms of Go were identified, Gi alpha 2 being predominant. As expected from a G protein-linked receptor, GTP and its nonhydrolyzable analogue GTP-gamma-S decreased tracer galanin binding to cell membranes. This resulted from a change in receptor affinity without any modification in the number of sites. Selective antibodies against the COOH-terminal decapeptide of the alpha-subunits of the Gi and Go proteins were used to block G protein interaction before we studied galanin binding. Antibody AS, which selectively recognizes Gi alpha 1 and Gi alpha 2, decreased tracer galanin binding to membranes at concentrations where there were no effects of other antibodies specifically directed against Gi alpha 3 or G alpha o. These data suggest that Gi1 and/or Gi2 interact with the galanin receptor and probably mediate the effects of galanin in pancreatic beta-cells.
Cormont
M
M
Unit 145, National Institute of Health and Medical Research, Faculty of Medicine, Nice, France.
Le Marchand-Brustel
Y
Y
Van Obberghen
E
E
Spiegel
A M
AM
Sharp
G W
GW
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D001921:Brain / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000032:analysis / Q000378:metabolism*; D019004:Galanin; D006150:Guanine Nucleotides / Q000494:pharmacology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D007118:Immunoassay; D007340:Insulinoma; D007668:Kidney / Q000378:metabolism; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009243:NAD / Q000378:metabolism; D010190:Pancreatic Neoplasms; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology / Q000378:metabolism; D037342:Pertussis Toxin; D051381:Rats; D044088:Receptors, Galanin; D011964:Receptors, Gastrointestinal Hormone / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D000906:Antibodies; D006150:Guanine Nucleotides; D046911:Macromolecular Substances; D010455:Peptides; D044088:Receptors, Galanin; D011964:Receptors, Gastrointestinal Hormone; D010566:Virulence Factors, Bordetella; D009243:NAD; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019004:Galanin; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins