Diacylglycerol kinase $zeta; regulates phosphatidylinositol 4-phosphate 5-kinase I$alpha; by a novel mechanism

Abstract

Phosphatidylinositol 4,5-bisphosphate (PIP2) plays an important role during actin polymerization and is produced by the type I phosphatidylinositol 4-phosphate 5-kinases (PIP5KI), which are activated by phosphatidic acid (PA). As diacylglycerol kinases (DGKs) generate PA by phosphorylating diacylglycerol (DAG), we investigated whether DGKs were involved in controlling PIP2 levels by regulating PIP5KI activity. Here we show that expression of DGKζ significantly enhances PIP5KIα activity in thrombin-stimulated HEK293 cells, and DGK activity is required for this stimulation. We also observed that DGKζ co-immunoprecipitated and co-localized with PIP5KIα, suggesting that they reside in a regulated signaling complex. To explore the role of DGKζ in actin polymerization, we examined the subcellular distribution of DGKζ, PIP5KIα and actin, and found that these proteins co-localized with actin in lamellipodial protrusions. Supporting that PIP5KIα regulation occurs at the sites of actin polymerization, we found that PIP2 also accumulated in the actin-rich regions of lamellipodia. Significantly, in wounding assays, DGKζ, PIP5KIα and PIP2 accumulated at the leading edge of migrating A172 cells, where massive actin polymerization is known to occur. Combined, these data support a novel function for DGKζ: by generating PA, it stimulates PIP5KIα activity to increase local PIP2, which regulates actin polymerization.