Functional involvement of VAMP/synaptobrevin-2 in cAMP-stimulated aquaporin 2 translocation in renal collecting duct cells.
The involvement of soluble N-ethylmaleimide sensitive factor-attachment protein receptor (SNARE) proteins in the cAMP-induced exocytosis of aquaporin 2 (AQP2)-containing vesicles was investigated in AQP2-transfected renal CD8 cells. RT-PCR and western blot analysis confirmed the presence of the SNARE homologs VAMP/synaptobrevin-2, syntaxin-1, syntaxin-4 and SNAP-23 in CD8 cells. Tetanus neurotoxin (TeNT) was efficient in cleaving synaptobrevin-like protein both in vitro and in intact CD8 cells incubated with the toxin. TeNT treatment in intact CD8 cells completely abolished cAMP-stimulated AQP2 targeting to the plasma membrane, as assessed by quantification of cell-surface immunoreactivity to an anti-AQP2 antibody raised against a peptide reproducing the extracellular AQP2 C-loop. These results represent the first evidence for the functional involvement of VAMP-2 in cAMP-induced AQP2 exocytosis in renal cells.
Dipartimento di Fisiologia Generale ed Ambientale, University of Bari, Via Amendola 165/A, 70126 Bari, Italy.
Sabine
S
Gouraud
Antonia
A
Laera
Giuseppe
G
Calamita
Monica
M
Carmosino
Giuseppe
G
Procino
Ornella
O
Rossetto
Roberta
R
Mannucci
Walter
W
Rosenthal
Maria
M
Svelto
Giovanna
G
Valenti
D000818:Animals; D000906:Antibodies; D051399:Aquaporin 2; D051403:Aquaporin 6; D020346:Aquaporins / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology*; D007150:Immunohistochemistry; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism*; D008565:Membrane Proteins / Q000187:drug effects / Q000378:metabolism*; D008666:Metalloendopeptidases; D008954:Models, Biological; D017434:Protein Structure, Tertiary / Q000502:physiology; D021381:Protein Transport / Q000502:physiology*; D050683:R-SNARE Proteins; D011817:Rabbits; D013744:Tetanus Toxin; D022161:Transport Vesicles / Q000378:metabolism*; D014882:Water-Electrolyte Balance / Q000502:physiology
D000906:Antibodies; D051399:Aquaporin 2; D051403:Aquaporin 6; D020346:Aquaporins; D008565:Membrane Proteins; D050683:R-SNARE Proteins; D013744:Tetanus Toxin; C035979:tetanospasmin; D000242:Cyclic AMP; D008666:Metalloendopeptidases