Abstract 10373: Characterization of WNK-PI3-Kinase Interaction and its Impact on Signaling by Both Enzymes
With no Lysine (WNKs) and Phosphoinostide 3-kinases (PI3Ks) are two enzyme families with central roles in the regulation of cellular functions and responses to various extracellular stimuli. Members of both enzyme families play essential roles in embryonic development, particularly in the cardiovascular system. We aim to characterize the functions and interacting partners of WNK family members, as these enzymes play important roles in various cellular processes and their signaling is disrupted in multiple disorders. WNKs have been implicated in blood pressure regulation via modulation of ion channels through the downstream effectors, SPAK and OSR. As such, mutations in WNK1 and WNK4 are the primary cause of PHA II, a hereditary form of hypertension. WNKs are large proteins (ranging from ~100-300 kDa) whose structures are largely uncharacterized, lacking distinct domains beyond the kinase domain. Since WNKs contain many proline-rich regions, we hypothesized a regulatory interaction between WNKs and the p85 regulatory subunit of class IA PI3Ks, which contains an SH3 domain. Surprisingly, we found a novel interaction between WNK1 and p85 involving not the SH3 but the phosphotyrosine-binding SH2 domain of p85. Given that p85 SH2 domains inhibit PI 3-kinase catalytic activity, this suggests a regulatory influence of WNK1 on the lipid kinase activity of p85/p110 heterodimers. We have defined the necessary residues on both interacting partners, showing that the kinase activity of WNK1 is not required for this interaction. Interestingly, our characterization of the WNK-PI3K complex has revealed an isoform-specific interaction of WNK1 with p85/p110β, suggesting possible implications for GPCR signaling to PI3Ks. Furthermore, our data suggest a crosstalk between p85/p110β binding to WNKs and signaling from either enzyme, evidenced by an altered cellular response to osmotic stress in p85 knockdown cells. Our study provides the first evidence of a direct interaction between WNKs and PI3Ks. These data provide novel insights into the regulation of both WNK and PI3K enzyme families, which may greatly alter our understanding of their functions and their roles in development and disease.
- © 2013 by American Heart Association, Inc.