Influence of tumor-promoting phorbol esters on the phosphorylation of membrane proteins in lymphocytes

Treatment of bovine lymphocytes with 12-O-tetradecanoyl-phorbol-13-acetate(TPA) (10^–8 M) for as little as 5 min significantly altersthe ability of membrane-particulate fractions to phosphorylateproteins using in situ protein kinases and exogenous [-³²P]ATP.After 20 min of treatment, the phosphorylation of two proteinswith mol. wts. of 65K and 74K in the isolated membrane-particulatefraction is increased approximately 30–35%, whereas thephosphorylation of a 130K protein is almost completely suppressed.The ability of different phorbol esters to alter membrane proteinphosphorylation correlates well with their potency as tumorpromoters in mouse epidermis and as comitogens in phytohemagglutinin-treatedlymphocytes. This phorbol ester response appears to have a lowtemperature coefficient since cells treated with TPA at 4?Calso responded, although at a slower rate. This action of TPAis neither mimicked nor antagonized by dibutyryl cAMP (1 mM);moreover, it is not affected by retinpic acid, an agent whichblocks several other phorbol ester effects in these cells. Inhibitionof protein synthesis with cycloheximide also fails to influencethis response. In contrast, trifluoperazine, an inhibitor ofcalmodulin function and certain lipid-dependent protein kinases,depressed the phosphorylation of the 65K and 74K proteins tobelow the control level.