Vivek Bhalla, MD

Publication Details

  • Phosphopeptide Screen Uncovers Novel Phosphorylation Sites of Nedd4-2 That Potentiate Its Inhibition of the Epithelial Na+ Channel JOURNAL OF BIOLOGICAL CHEMISTRY Hallows, K. R., Bhalla, V., Oyster, N. M., Wijngaarden, M. A., Lee, J. K., Li, H., Chandran, S., Xia, X., Huang, Z., Chalkley, R. J., Burlingame, A. L., Pearce, D. 2010; 285 (28): 21671-21678

    Abstract:

    The E3 ubiquitin ligase Nedd4-2 regulates several ion transport proteins, including the epithelial Na(+) channel (ENaC). Nedd4-2 decreases apical membrane expression and activity of ENaC. Although it is subject to tight hormonal control, the mechanistic basis of Nedd4-2 regulation remains poorly understood. To characterize regulatory inputs to Nedd4-2 function, we screened for novel sites of Nedd4-2 phosphorylation using tandem mass spectrometry. Three of seven identified Xenopus Nedd4-2 Ser/Thr phosphorylation sites corresponded to previously identified target sites for SGK1, whereas four were novel, including Ser-293, which matched the consensus for a MAPK target sequence. Further in vitro and in vivo phosphorylation experiments revealed that Nedd4-2 serves as a target of JNK1, but not of p38 MAPK or ERK1/2. Additional rounds of tandem mass spectrometry identified two other phosphorylated residues within Nedd4-2, including Thr-899, which is present within the catalytic domain. Nedd4-2 with mutations at these sites had markedly inhibited JNK1-dependent phosphorylation, virtually no ENaC inhibitory activity, and significantly reduced ubiquitin ligase activity. These data identify phosphorylatable residues that activate Nedd4-2 and may work together with residues targeted by inhibitory kinases (e.g. SGK1 and protein kinase A) to govern Nedd4-2 regulation of epithelial ion transport.

    View details for DOI 10.1074/jbc.M109.084731

    View details for Web of Science ID 000279516100054

    View details for PubMedID 20466724

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