Subtype-specific translocation of diacylglycerol kinase α and γ and its correlation with protein kinase C

Abstract

We examined the translocation of diacylglycerol kinase (DGK) α and γ fused with green fluorescent protein in living Chinese hamster ovary K1 cells (CHO-K1) and investigated temporal and spatial correlations between DGK and protein kinase C (PKC) when both kinases are overexpressed. DGKα and γ were present throughout the cytoplasm of CHO-K1 cells. Tetradecanoylphorbol 13-acetate (TPA) induced irreversible translocation of DGKγ, but not DGKα, from the cytoplasm to the plasma membrane. The (TPA)-induced translocation of DGKγ was inhibited by the mutation of C1A but not C1B domain of DGKγ and was not inhibited by staurosporine. Arachidonic acid induced reversible translocation of DGKγ from the cytoplasm to the plasma membrane, whereas DGKα showed irreversible translocation to the plasma membrane and the Golgi network. Purinergic stimulation induced reversible translocation of both DGKγ and α to the plasma membrane. The timing of the ATP-induced translocation of DGKγ roughly coincided with that of PKCγ re-translocation from the membrane to the cytoplasm. Furthermore, re-translocation of PKCγ was obviously hastened by co-expression with DGKγ and was blocked by an inhibitor of DGK (R59022). These results indicate that DGK shows subtype-specific translocation depending on extracellular signals and suggest that PKC and DGK are orchestrated temporally and spatially in the signal transduction.