Phorbol Esters Mediate Phospholipid-free Activation of Rat Brain Protein Kinase C1

The present study provides evidence that rat brain protein kinase C elicits a phosphotransferase activity towards histone and undergoes autophosphorylation in the absence of phosphatidylserine. The tumor pro moter 12-0-tetradecanoylphorbol-13-acetate binds to and activates pro tein kinase C in a phospholipid-free reaction. The apparent activation constant (A1,, = 2.7 IIM) is not modified by the absence of phospholipid but the maximum velocity is greatly decreased. The phosphotransfer reaction to exogenous substrates occurs in 0.5 HIM ethylenebis(oxyethylenenitrilo)tetraacetic acid, although autophosphorylation in these conditions requires the presence of ( a'*. The protein kinase C inhibitor (l-(5-isoquinolinesulfonyl)-2-methylpiperazine inhibits the re action, whereas the cAMP-dependent protein kinase inhibitor is ineffec tive. In contrast to diacylglycerol, which is a poor activator, unsaturated fatty acids potently activate the phospholipid-free reaction. Moreover, the substrate specificity is markedly changed, e.g., myelin basic protein and histone types VI-S and VII-S appear to be relatively better substrates in the phospholipid-free reaction. The data presented indicate that protein kinase C (or some individual isoforms) may function, at least partially, without binding to membrane phospholipid and suggest that this novel characteristic of phorbol esters may account for their tumor-promoting activity.