Small-molecule screening identifies modulators of aquaporin-2 trafficking

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Item Type:	Article
Title:	Small-molecule screening identifies modulators of aquaporin-2 trafficking
Creators Name:	Bogum, J., Faust, D., Zühlke, K., Eichhorst, J., Moutty, M.C., Furkert, J., Eldahshan, A., Neuenschwander, M., von Kries, J.P., Wiesner, B., Trimpert, C., Deen, P.M., Valenti, G., Rosenthal, W. and Klussmann, E.
Abstract:	In the principal cells of the renal collecting duct, arginine vasopressin (AVP) stimulates the synthesis of cAMP, leading to signaling events that culminate in the phosphorylation of aquaporin-2 water channels and their redistribution from intracellular domains to the plasma membrane via vesicular trafficking. The molecular mechanisms that control aquaporin-2 trafficking and the consequent water reabsorption, however, are not completely understood. Here, we used a cell-based assay and automated immunofluorescence microscopy to screen 17,700 small molecules for inhibitors of the cAMP-dependent redistribution of aquaporin-2. This approach identified 17 inhibitors, including 4-acetyldiphyllin, a selective blocker of vacuolar H(+)-ATPase that increases the pH of intracellular vesicles and causes accumulation of aquaporin-2 in the Golgi compartment. Although 4-acetyldiphyllin did not inhibit forskolin-induced increases in cAMP formation and downstream activation of protein kinase A (PKA), it did prevent cAMP/PKA-dependent phosphorylation at serine 256 of aquaporin-2, which triggers the redistribution to the plasma membrane. It did not, however, prevent cAMP-induced changes to the phosphorylation status at serines 261 or 269. Last, we identified the fungicide fluconazole as an inhibitor of cAMP-mediated redistribution of aquaporin-2, but its target in this pathway remains unknown. In conclusion, our screening approach provides a method to begin dissecting molecular mechanisms underlying AVP-mediated water reabsorption, evidenced by our identification of 4-acetyldiphyllin as a modulator of aquaporin-2 trafficking.
Keywords:	Aquaporin 2, Benzodioxoles, Benzofurans, Cultured Cells, Cyclic AMP, Fluconazole, Fluorescence Microscopy, Forskolin, Golgi Apparatus, High-Throughput Screening Assays, Phosphorylation, Protein Transport, Vacuolar Proton-Translocating ATPases, Animals, Mice, Rats
Source:	Journal of the American Society of Nephrology
ISSN:	1046-6673
Publisher:	American Society of Nephrology
Volume:	24
Number:	5
Page Range:	744-758
Date:	April 2013
Official Publication:	https://doi.org/10.1681/ASN.2012030295
PubMed:	View item in PubMed

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