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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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