CCK, carbachol, and bombesin activate distinct PLC-beta isoenzymes via Gq/11 in rat pancreatic acinar membranes.
Four different isoforms of phospholipase C-beta (PLC-beta 1-4) have been discovered, raising the important question of whether a distinct receptor activates a single PLC-beta isoenzyme or a subset of PLC-beta isoenzymes. The present study was designed to investigate activation of PLC-beta isoenzymes by three different PLC-activating agonists that bind to different receptor entities, i.e., cholecystokinin octapeptide (CCK-8), bombesin, and carbachol in rat pancreatic acinar membranes. PLC activity was measured using exogenous [3H]phosphatidylinositol 4,5-bisphosphate as substrate. Western blot analysis of pancreatic acinar membranes revealed the presence of PLC-beta 1, -beta 3, -gamma 1, and -delta 1, but not of PLC-beta 2, -beta 4, -gamma 2, and -delta 2. Preincubation of the membranes with anti-PLC-beta 1 or -beta 3 antibody reduced agonist-induced activation of PLC. The order of sensitivity toward inhibition by anti-PLC-beta 1 antibody was CCK-8 > bombesin > carbachol. An opposite order of sensitivity was found for inhibition of PLC activity by anti-PLC-beta 3 antibody (carbachol > bombesin > CCK-8). Anti-PLC-beta 2, -beta 4, -gamma 1, -gamma 2, -delta 1, and -delta 2 antibodies had no effect. Preincubation of the membranes with an antibody raised against the COOH terminus of the alpha-subunit of Gq/11 proteins inhibited PLC activity in response to all three different receptor agonists to a similar extent, whereas anti-Gi alpha 1-2 and anti-Gi alpha 3 antibodies had no effect. In conclusion, the data of the present study indicate that CCK-8 and carbachol activate PLC-beta 1 and PLC-beta 3, respectively, whereas bombesin activates both PLC-beta 1 and PLC-beta 3. Activation of PLC-beta by these receptor agonists is mediated by Gq/11.
Piiper
A
A
II. Medical Department, University of Frankfurt, Germany.
Stryjek-Kaminska
D
D
Klengel
R
R
Zeuzem
S
S
D000818:Animals; D000906:Antibodies / Q000276:immunology; D001839:Bombesin / Q000494:pharmacology*; D002217:Carbachol / Q000494:pharmacology*; D002462:Cell Membrane / Q000378:metabolism; D002766:Cholecystokinin / Q000494:pharmacology*; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000276:immunology / Q000502:physiology*; D007527:Isoenzymes / Q000378:metabolism*; D010179:Pancreas / Q000378:metabolism*; D051381:Rats; D010738:Type C Phospholipases / Q000276:immunology / Q000378:metabolism*
D000906:Antibodies; D007527:Isoenzymes; D002217:Carbachol; D002766:Cholecystokinin; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins; D001839:Bombesin