Expression of SNARE proteins in enteroendocrine cell lines and functional role of tetanus toxin-sensitive proteins in cholecystokinin release.
In neurons, synaptic vesicle exocytosis involves the formation of a core complex particle including syntaxin-1, synaptosomal-associated protein of 25 kDa (SNAP-25) and vesicle-associated membrane protein (VAMP)-2/synaptobrevin. The expression of these proteins was investigated in a panel of cell lines, including lines of endocrine and intestinal origin, by Western blotting and/or immunocytochemistry. The three core complex proteins were detected in the enteroendocrine, cholecystokinin (CCK)-secreting, cell lines STC-1 and GLUTag, and in the endocrine non-intestinal cell lines CA-77 and HIT-T15. In contrast, SNAP-25 and syntaxin-1 were undetected in the intestinal non-endocrine cell lines IEC-6, HT-29 and Caco-2, whereas a slight expression of VAMP-2 was documented in IEC-6 and HT-29 cells. Co-immunoprecipitation experiments indicated that syntaxin-1, SNAP-25 and VAMP-2 were present in a complex similar to that identified in brain. In the STC-1 cell line, treatment of streptolysin-O-permeabilized cells with tetanus toxin (Tetx) selectively cleaved VAMP-2 and VAMP-3/cellubrevin, and simultaneously abolished Ca2+-induced CCK secretion (IC50 approximately 12 nM). These results show that endocrine cell lines of intestinal origin express syntaxin-1, SNAP-25 and VAMP-2, and suggest a key role for a Tetx-sensitive protein (for example VAMP-2 and/or VAMP-3) in the CCK secretion by STC-1 cells.
Némoz-Gaillard
E
E
INSERM Unité 45, Hôpital Edouard-Herriot, Lyon, France.
Bosshard
A
A
Regazzi
R
R
Bernard
C
C
Cuber
J C
JC
Takahashi
M
M
Catsicas
S
S
Chayvialle
J A
JA
Abello
J
J
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002766:Cholecystokinin / Q000378:metabolism*; D006224:Cricetinae; D004702:Endocrine Glands / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D007413:Intestinal Mucosa / Q000378:metabolism*; D007422:Intestines / Q000166:cytology; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D051381:Rats; D013744:Tetanus Toxin / Q000494:pharmacology*
D008565:Membrane Proteins; D013744:Tetanus Toxin; D002766:Cholecystokinin; D002118:Calcium