ACTIVATION OF TYPE-D PHOSPHOLIPASE BY SERUM STIMULATION AND RAS-INDUCED TRANSFORMATION IN NIH3T3 CELLS

Mitogenic stimulation of NIH3T3 fibroblasts with growth factors or ras oncogenes is associated with an increase in the levels of phosphorylc holine and diacylglycerol. Both metabolites could be generated as a re sult of direct activation of a phosphatidylcholine-specific phospholip ase C (PC-PLC) or by a more complex pathway, involving activation of p hospholipase D followed by choline kinase and phosphatidic acid-hydrol ase. We show evidence indicating that the generation of phosphorylchol ine and diacylglycerol follow independent mechanisms in both serum-tre ated and in ras-transformed NIH3T3 cells. No significant activation of a PC-PLC enzyme was observed. Instead, activation of a phosphatidylch oline-specific phospholipase D (PC-PLD) was detected. Moreover, while a fivefold constitutive activation of the endogenous PLD activity and a twofold increase on the levels of phosphatidic acid were observed in uas-transformed cells, very small alterations on these parameters wer e detected at late times after serum stimulation of quiescent cells. T hus, cell proliferation induced by uas oncogenes in fibroblasts cells may be functionally linked to activation of a PC-PLD enzyme. The diffe rences found in the activation of this enzyme between ras-transformed and normal cells may constitute an important difference in mitogenic s ignalling between normal and transformed cells.