Functional Involvement of VAMP/Synaptobrevin-2 in cAMP-Stimulated Aquaporin 2 Translocation in Renal Collecting Duct Cells

Title: Functional Involvement of VAMP/Synaptobrevin-2 in cAMP-Stimulated Aquaporin 2 Translocation in Renal Collecting Duct Cells
Authors: Gouraud, Sabine; Laera, Antonia; Calamita, Giuseppe; Carmosino, Monica; Procino, Giuseppe; Rossetto, Ornella; Mannucci, Roberta; Rosenthal, Walter; Svelto, Maria; Valenti, Giovanna
Publisher: Journal of Cell Science
Date Published: September 15, 2002
Reference Number: 576
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.
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This translation by the NDI Foundation is to assist the lay reader. To provide a clear, accessible interpretation of the original article, we eliminated or simplified some technical detail and complicated scientific language. We concentrated our translation on those aspects of the article dealing directly with NDI. The NDI Foundation thanks the researchers for their work toward understanding and more effectively treating this disorder.
© Copyright NDI Foundation 2007 (JC)

The protein, aquaporin-2 (AQP2) is found in the principal cells of the kidney collecting duct. It rests in the cell interior inside little sacs called vesicles. When signaled by a molecular sequence initiated by the hormone, vasopressin, the vesicles containing AQP2 travel from the cell interior to a specific section of the cell membrane. When they arrive there, AQP2 is inserted into the membrane where it becomes a channel through which water crosses through the membrane and enters the cell.

The AQP2 containing vesicles cannot make this trip to the membrane by themselves. They are helped by other proteins, not all of which have been determined. Gouraud, et al., suspected that a type of soluble N-ethymaleimide sensitive factor-attachment protein receptor (SNARE), namely VAMP-2, was involved in the AQP2 vesicle movement to the cell membrane.

To test their hypothesis, they generated laboratory cell cultures that contained AQP2s and VAMP-2s. When stimulated with cAMP, the AQP2s made their journey to the membrane. The researchers then added tetanus neurotoxins (TeNTs) to these cultures. TeNT is known to split VAMP-2 and thus interfere with its ability to function. After TeNT was injected into the cell cultures, they were again stimulated with cAMP. The result was that there was no travel of AQP2 laden vesicles to the cell membrane. This result provides evidence that VAMP-2 is directly involved in the cAMP-2 stimulated movement of AQP2 from the cell interior to cell membrane.