Protein kinase C inhibits binding of diacylglycerol kinase-zeta 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-zeta (DGKzeta) to phosphorylate the lipid second messenger diacylglycerol into phosphatidic acid. pRB binds to the MARCKS phosphorylation-site domain of DGKzeta that can be phosphorylated by protein kinase C (PKC). Here, we report that activation of PKC by phorbol ester inhibits DGKzeta 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 DGKzeta-MARCKS phosphorylation-site domain also prevented DGKzeta binding to pRB, suggesting that PKC phosphorylation of these residues negatively regulates the interaction between DGKzeta and pRB. In PKC overexpression studies, it appeared that activation of particularly the (wild-type) PKCalpha isoform inhibits DGKzeta binding to pRB, whereas dominant-negative PKCalpha neutralized this inhibition. PKCalpha activation thus prevents DGKzeta regulation by pRB, which may have implications for nuclear diacylglycerol and phosphatidic acid levels during the cell cycle.
Los
Alrik P
AP
Division of Cellular Biochemistry, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands.
de Widt
John
J
Topham
Matthew K
MK
van Blitterswijk
Wim J
WJ
Divecha
Nullin
N
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D019852:Diacylglycerol Kinase / Q000737:chemistry / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D007527:Isoenzymes / Q000145:classification / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D010768:Phosphoserine / Q000378:metabolism; D011485:Protein Binding; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism*
D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; C000619710:MARCKS protein, human; D008565:Membrane Proteins; D016160:Retinoblastoma Protein; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D010768:Phosphoserine; D019852:Diacylglycerol Kinase; D011493:Protein Kinase C