Actions of Rab3 effector domain peptides in chief cells from guinea pig stomach.
Rab3 proteins are low molecular weight guanine nucleotide-binding proteins that belong to the Ras superfamily and are believed to play a role in the final steps of exocytosis. To examine potential interactions of these proteins with signaling pathways that mediate pepsinogen secretion from gastric chief cells, we synthesized peptides corresponding to the effector domain of Rab3. In the absence of added calcium [calcium concentration ([Ca2+]) < 1 nM], a maximal concentration (15 microM) of the Rab3 effector domain peptide or Rab3AL peptide, containing alanine and leucine substitutions, stimulated the release of 62 and 66%, respectively, of total pepsinogen from streptolysin O-permeabilized chief cells. A Rab2AL peptide, corresponding to the Rab2 effector domain, and modified (scrambled and truncated) Rab3AL peptides did not alter secretion from permeabilized cells. An additive secretory response was observed when 5 microM Rab3AL peptide was combined with increasing calcium ([Ca2+] < 1 nM to 3 microM). In contrast, adding up to 3 mM adenosine 3',5'-cyclic monophosphate (cAMP) had no effect on Rab3AL peptide-induced secretion, and Rab3AL peptide did not alter endogenous cAMP production. The addition of a nonhydrolyzable GTP analogue [0.01 to 100 microM guanosine 5'-O-(3-thiotriphosphate)] potentiated the secretory response to Rab3AL peptide. This potentiated response indicates that other GTP-binding proteins are involved in calcium-independent secretion. Preincubation of cells with streptolysin O (10-30 min), to allow egress of cytosolic constituents, enhanced the response to Rab3AL peptide, suggesting that the target(s) for this peptide is (are) anchored to chief cell membranes.(ABSTRACT TRUNCATED AT 250 WORDS)
Singh
G
G
Department of Medicine, State University of New York Health Science Center, Brooklyn 11203-2098, USA.
Raffaniello
R D
RD
Eng
J
J
Raufman
J P
JP
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D000242:Cyclic AMP / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D019204:GTP-Binding Proteins / Q000737:chemistry*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006168:Guinea Pigs; D008297:Male; D008969:Molecular Sequence Data; D010435:Pepsinogens / Q000378:metabolism; D010446:Peptide Fragments / Q000494:pharmacology*; D013270:Stomach / Q000166:cytology / Q000187:drug effects* / Q000648:ultrastructure; D020692:rab3 GTP-Binding Proteins
D010435:Pepsinogens; D010446:Peptide Fragments; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000242:Cyclic AMP; D019204:GTP-Binding Proteins; D020692:rab3 GTP-Binding Proteins; D002118:Calcium