Vesicle-associated membrane protein 2 is essential for cAMP-regulated exocytosis in rat parotid acinar cells. The inhibition of cAMP-dependent amylase release by botulinum neurotoxin B.
Amylase exocytosis of the parotid gland is mediated by intracellular cAMP. To investigate whether cAMP-dependent secretion has a mechanism similar to that of regulated neuroexocytosis, we examined the expression of synaptosome-associated proteins. In rat parotid acinar cells, we found 25 (p25) and 18 kDa (p18) proteins reacted with antibodies against Rab3A and vesicle-associated membrane protein 2 (VAMP-2), respectively. On the other hand, syntaxin 1 and SNAP-25, which interact with VAMP-2 at synapses, were undetectable. Rab3A-like p25 and VAMP-2-like p18 were also expressed in other exocrine acinar cells. The latter was localized at secretory granule membranes, and the former was detected in secretory granule and cytosolic fractions. The antibody against VAMP-2 used in this study did not react with cellubrevin, and p18 was cleaved with botulinum neurotoxin B. Thus, we identified p18 as VAMP-2. Botulinum neurotoxin B inhibited the cAMP-induced amylase release from streptolysin O-permeabilized acinar cells. Therefore, VAMP-2 is required for cAMP-regulated amylase release in rat parotid acinar cells. This is the first report that VAMP-2 is involved in regulated exocytosis that is independent of Ca2+.
Fujita-Yoshigaki
J
J
Department of Physiology, Nihon University School of Dentistry at Matsudo, 2-870-1 Sakae-cho Nishi, Matsudo, Chiba 271, Japan.
Dohke
Y
Y
Hara-Yokoyama
M
M
Kamata
Y
Y
Kozaki
S
S
Furuyama
S
S
Sugiya
H
H
D000681:Amylases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000818:Animals; D001905:Botulinum Toxins / Q000494:pharmacology*; D002462:Cell Membrane / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D005089:Exocytosis; D019204:GTP-Binding Proteins / Q000378:metabolism; D006868:Hydrolysis; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010306:Parotid Gland / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D050683:R-SNARE Proteins; D051381:Rats; D017207:Rats, Sprague-Dawley; D020692:rab3 GTP-Binding Proteins
D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D050683:R-SNARE Proteins; D000242:Cyclic AMP; D000681:Amylases; D001905:Botulinum Toxins; D019204:GTP-Binding Proteins; D020692:rab3 GTP-Binding Proteins