ALTERATION OF GLYCEROLIPID AND SPHINGOLIPID-DERIVED 2ND-MESSENGER KINETICS IN RAS TRANSFORMED 3T3 CELLS

The effect of ras transformation (rasB fibroblasts) on basal and serum -stimulated diacylglycerol (DAG) composition and mass was examined ove r time with respect to changes in membrane phospholipid composition an d ceramide mass. RasB cells vs. nontransformed control cells (rasD and NR6) had chronically elevated DAG levels (up to 240 min) following se rum stimulation, indicating a defect in the recovery phase of the intr acellular DAG pulse. Ras transformation also had a dramatic effect on DAG composition, Molecular species analysis revealed that DAG from uns timulated rasB cells was enriched in the Delta 9 desaturase fatty acyl species (monoenoate 18:1(n - 7) and 18:1(n - 9)), acid depleted in ar achidonic acid (20:4(n - 6)). With the exception of glycerophosphoinos itol (GPI), DAG remodeling paralleled the compositional alterations in individual phospholipid classes. Importantly, ras transformation alte red the fatty acyl composition of sphingomyelin, a precursor to the ce ramide second messenger. With the addition of serum, control cells (ra sD) had a progressive increase in ceramide mass with levels approximat ely 5-fold higher by 240 min. In contrast, ceramide levels did not inc rease in rasB cells at either 4 or 240 min. These results demonstrate that ras-oncogene, in addition to its effects on DAG metabolism, can a lso abolish the cellular increase in ceramide mass in response to seru m stimulation. Since DAG and ceramide may have opposing biological fun ctions, the prolonged elevation of DAG and the suppression of ceramide levels would be consistent with an enhanced proliferative capacity.