Effect of a Rab3A effector domain-related peptide, CCK, and EGF in permeabilized pancreatic acini.
We report here that a synthetic peptide of the effector domain of the small-molecular-weight GTP-binding protein Rab3A (EDRab3AL) is a potent stimulator of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] production and amylase secretion in digitonin-permeabilized pancreatic acini. Moreover, the Rab3A effector domain peptide caused phosphatidylinositol 4,5-bisphosphate breakdown, indicating that the observed increase in Ins(1,4,5)P3 is due to stimulation of a phosphoinositide-specific phospholipase C (PLC). The dose-response curve for EDRab3AL-induced amylase release was biphasic, showing a maximum at 0.3 nM EDRab3AL and a decline at higher peptide concentrations. By contrast, the dose-response curve for EDRab3AL-induced Ins(1,4,5)P3 production was monophasic, showing stimulation with increasing EDRab3AL concentrations. A peptide of the effector domain of Rab1A, EDRab1AL, had no effect, indicating that the response to EDRab3AL is specific. Cholecystokinin octapeptide (CCK-8) and EDRab3AL had additive effects on the acinar Ins(1,4,5)P3 level. Epidermal growth factor (EGF), which has recently been shown to inhibit CCK-8-induced Ins(1,4,5)P3 production in pancreatic acinar cells, also decreased EDRab3AL-induced Ins(1,4,5)P3 production. These results suggest that EDRab3AL and CCK-8 act on the same EGF-inhibitable PLC by independent mechanisms. CCK-8 increased and EGF decreased amylase release in response to submaximal EDRab3AL concentrations. By contrast, at supramaximal EDRab3AL concentrations EGF increased and CCK-8 decreased EDRab3AL-stimulated amylase release. EDRab3AL had no effect in intact acini, indicating that the site of action of EDRab3AL is intracellular. We conclude that EDRab3AL regulates phosphoinositide-specific PLC activity and thereby amylase secretion in an analogous fashion to CCK-8, but from within the cell.(ABSTRACT TRUNCATED AT 250 WORDS)
Zeuzem
S
S
Department of Internal Medicine, University of Frankfurt, Germany.
Stryjek-Kaminska
D
D
Caspary
W F
WF
Stein
J
J
Piiper
A
A
D000681:Amylases / Q000378:metabolism; D000818:Animals; D000242:Cyclic AMP / Q000096:biosynthesis; D004072:Digitonin / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000737:chemistry*; D006868:Hydrolysis; D015544:Inositol 1,4,5-Trisphosphate / Q000096:biosynthesis; D008297:Male; D010179:Pancreas / Q000187:drug effects* / Q000378:metabolism; D010446:Peptide Fragments / Q000494:pharmacology*; D010539:Permeability; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012844:Sincalide / Q000494:pharmacology*
D010446:Peptide Fragments; D004815:Epidermal Growth Factor; D015544:Inositol 1,4,5-Trisphosphate; D000242:Cyclic AMP; D017853:Phosphotransferases (Alcohol Group Acceptor); C068961:phosphatidylinositol 4,5-biphosphate kinase; D000681:Amylases; D019204:GTP-Binding Proteins; D004072:Digitonin; D012844:Sincalide