Phorbol ester inhibits the phosphorylation of the retinoblastoma protein without suppressing cyclin D-associated kinase in vascular smooth muscle cells.
To elucidate the role of protein kinase C in vascular smooth muscle cell proliferation, we examined the effects of phorbol 12-myristate 13-acetate (PMA) on G1 events in human arterial cells. About 15 h after G0 cells were stimulated with fetal bovine serum and basic fibroblast growth factor, [3H]thymidine incorporation started. PMA (10 nM) inhibited the incorporation over 90% when added earlier than 3 h after stimulation, but had no effect when added 12 h or later. PMA inhibited the phosphorylation of the retinoblastoma protein (pRb), which normally began at about 9 h. PMA did not inhibit the gene expression of Cdk2, Cdk3, Cdk4, Cdk5, and cyclins G, C, and D, all of which began at 0-3 h. However, PMA reduced the expression of cyclins E and A, which usually began at 3-9 h and about 15 h, respectively. PMA inhibited the histone H1 kinase activity of Cdk2, which increased from about 9 h, whereas PMA did not inhibit the pRb kinase activities of cyclin D-associated kinase(s) and Cdk4, detectable from 0-3 h. These results suggested that the PMA-induced inhibition of pRb phosphorylation is not mediated by suppressing cyclin D-associated kinase(s) including Cdk4, but involves the suppression of Cdk2 activity that results from the reduced expression of cyclins E and A.
Sasaguri
T
T
National Cardiovascular Center Research Institute, Osaka University, Suita, Osaka 565, Japan.
Ishida
A
A
Kosaka
C
C
Nojima
H
H
Ogata
J
J
D002453:Cell Cycle / Q000187:drug effects*; D002478:Cells, Cultured; D056741:Cyclin D; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D010703:Phorbol Esters / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000502:physiology*; D011518:Proto-Oncogene Proteins; D016160:Retinoblastoma Protein / Q000378:metabolism*
D056741:Cyclin D; D016213:Cyclins; D010703:Phorbol Esters; D011518:Proto-Oncogene Proteins; D016160:Retinoblastoma Protein; D011493:Protein Kinase C; C495900:CDK4 protein, human; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases