Protein kinase C inhibits binding of diacylglycerol kinase-ζ to the retinoblastoma protein

We previously showed that the retinoblastoma protein (pRB), a key regulator of G1 to S-phase transition of the cell cycle, binds to and stimulates diacylglycerol kinase-ζ (DGKζ) to phosphorylate the lipid second messenger diacylglycerol into phosphatidic acid. pRB binds to the MARCKS phosphorylation-site domain of DGKζ that can be phosphorylated by protein kinase C (PKC). Here, we report that activation of PKC by phorbol ester inhibits DGKζ binding to pRB. Ro 31-8220, a specific inhibitor of PKC, alleviated this inhibition of binding. Mimicking of PKC phosphorylation of serine residues (by S/D but not S/N mutations) within the DGKζ-MARCKS phosphorylation-site domain also prevented DGKζ binding to pRB, suggesting that PKC phosphorylation of these residues negatively regulates the interaction between DGKζ and pRB. In PKC overexpression studies, it appeared that activation of particularly the (wild-type) PKCα isoform inhibits DGKζ binding to pRB, whereas dominant-negative PKCα neutralized this inhibition. PKCα activation thus prevents DGKζ regulation by pRB, which may have implications for nuclear diacylglycerol and phosphatidic acid levels during the cell cycle.