WNK1 promotes PIP₂ synthesis to coordinate growth factor and GPCR-G_q signaling

Background: PLC-β signaling is generally thought to be mediated by allosteric activation by G proteins and Ca²⁺. Although availability of the phosphatidylinositol-4,5-biphosphate (PIP₂) substrate is limiting in some cases, its production has not been shown to be independently regulated as a signaling mechanism. WNK1 protein kinase is known to regulate ion homeostasis and cause hypertension when expression is increased by gene mutations. However, its signaling functions remain largely elusive. Results: Using diacylglycerol-stimulated TRPC6 and inositol trisphosphate-mediated Ca²⁺ transients as cellular biosensors, we show that WNK1 stimulates PLC-β signaling in cells by promoting the synthesis of PIP₂ via stimulation of phosphatidylinositol 4-kinase IIIα. WNK1 kinase activity is not required. Stimulation of PLC-β by WNK1 and by Gα_q are synergistic; WNK1 activity is essential for regulation of PLC-β signaling by G_q-coupled receptors, and basal input from G_q is necessary for WNK1 signaling via PLC-β. WNK1 further amplifies PLC-β signaling when it is phosphorylated by Akt kinase in response to insulin-like growth factor. Conclusions: WNK1 is a novel regulator of PLC-β that acts by controlling substrate availability. WNK1 thereby coordinates signaling between G protein and Akt kinase pathways. Because PIP₂ is itself a signaling molecule, regulation of PIP₂ synthesis by WNK1 also allows the cell to initiate PLC signaling while independently controlling the effects of PIP₂ on other targets. These findings describe a new signaling pathway for Akt-activating growth factors, a mechanism for G protein-growth factor crosstalk, and a means to independently control PLC signaling and PIP₂ availability.