Evidence of heterotrimeric G-protein involvement in regulated exocytosis from permeabilized pancreatic acini.
Constitutive membrane trafficking events are regulated by heterotrimeric G-proteins (G-proteins) in addition to their regulation by small GTP-binding proteins (smgs). Here, we used streptolysin O-permeabilized mouse pancreatic acini and compounds that interact with G-proteins, but not smgs, to examine whether G-proteins are also involved in regulated pancreatic exocytosis. The wasp venom mastoparan (10 microM) inhibited by 25-50% amylase release from permeabilized acini stimulated by various combinations of Ca2+, cyclic AMP (cAMP), 12-O-tetradecanoylphorbol 13-acetate, and guanosine (5'-[gamma-thio]triphosphate (GTP gamma S), while the inactive analogue Mas17 was without effect. Pretreatment of intact acini with pertussis toxin resulted in an approximately 30% reduction of amylase secretion from cells subsequently permeabilized and stimulated with calcium and GTP gamma S. Pretreatment of intact acini with cholera toxin increased stimulated amylase release by 30% from subsequently permeabilized cells, and this effect was mimicked by 8-Br-cAMP. The cAMP-dependent protein kinase inhibitor H-89 (3 microM) largely reversed the effect of cholera toxin, indicating that cholera toxin's effect is due to increased cellular cAMP levels. The inhibitory effects of mastoparan and pertussis toxin suggest that a Gi/Go-type G-protein(s) is (are) involved in the regulation of exocytosis. Since mastoparan inhibited exocytosis stimulated by all intracellular mediators tested, it indicates that the G-protein acts at a distal step in the exocytic process.
De Lisle
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RC
Department of Anatomy and Cell Biology, University of Kansas Medical Center, Kansas City 66160, USA.
Howell
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GW
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000818:Animals; D001426:Bacterial Proteins; D002118:Calcium / Q000494:pharmacology; D002463:Cell Membrane Permeability; D002772:Cholera Toxin / Q000494:pharmacology; D000242:Cyclic AMP / Q000494:pharmacology; D005089:Exocytosis; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000502:physiology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D036341:Intercellular Signaling Peptides and Proteins; D007546:Isoquinolines / Q000494:pharmacology; D046911:Macromolecular Substances; D051379:Mice; D010179:Pancreas / Q000378:metabolism*; D010455:Peptides; D037342:Pertussis Toxin; D047428:Protein Kinase Inhibitors; D013301:Streptolysins / Q000494:pharmacology; D013449:Sulfonamides; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D014862:Wasp Venoms / Q000494:pharmacology
D001426:Bacterial Proteins; D036341:Intercellular Signaling Peptides and Proteins; D007546:Isoquinolines; D046911:Macromolecular Substances; D010455:Peptides; D047428:Protein Kinase Inhibitors; D013301:Streptolysins; D013449:Sulfonamides; D010566:Virulence Factors, Bordetella; D014862:Wasp Venoms; C025964:streptolysin O; D015124:8-Bromo Cyclic Adenosine Monophosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C037500:mastoparan; D002772:Cholera Toxin; D000242:Cyclic AMP; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium