EPIDERMAL GROWTH-FACTOR INDUCES THE PRODUCTION OF BIOLOGICALLY DISTINGUISHABLE DIGLYCERIDE SPECIES FROM PHOSPHATIDYLINOSITOL AND PHOSPHATIDYLCHOLINE VIA THE INDEPENDENT ACTIVATION OF TYPE-C AND TYPE-D PHOSPHOLIPASES

An early response to epidermal growth factor in A431 cells is the gene ration of diglyceride, a physiological activator of protein kinase C. By differentially prelabeling cellular phospholipids with [H-3]arachid onate and [H-3]myristate, which are incorporated primarily into phosph atidylinositol and phosphatidylcholine, respectively, we have found th at epidermal growth factor induces an increase in diglyceride levels f rom both phosphatidylinositol and phosphatidylcholine via distinct mec hanisms and kinetics. The epidermal growth factor-induced increase in phosphatidylinositol-derived diglyceride was transient and peaked at 5 min. As diglyceride levels dropped, there was a corresponding increas e in phosphatidic acid, suggesting that the diglyceride is efficiently converted to phosphatidic acid by a diglyceride kinase. In contrast, epidermal growth factor-induced increases in phosphatidylcholine-deriv ed diglyceride peaked at 30 min and remained elevated for greater than 2 h. The epidermal growth factor-induced increases in phosphatidic ac id detected in [H-3]myristate-prelabeled cells paralleled the increase in diglyceride, suggesting that the phosphatidylcholine-derived digly ceride is produced from phosphatidic acid via a phosphatidic acid phos phatase. Consistent with this hypothesis, epidermal growth factor also induced a protein kinase C-independent phospholipase D activity that was specific for phosphatidylcholine. These data suggest that epiderma l growth factor induces diglyceride production from phosphatidylinosit ol and phosphatidylcholine via two distinct mechanisms: a rapid and tr ansient induction of diglyceride that likely involves phospholipase C- gamma-mediated hydrolysis of phosphatidylinositol-4,5-bisphosphate and a slower, more sustained induction of diglyceride via a phospholipase D-mediated hydrolysis of phosphatidylcholine to produce phosphatidic acid, which is then converted to diglyceride by a phosphatidic acid ph osphatase. Since the diglycerides produced from phosphatidylinositol a nd phosphatidylcholine are differentially metabolized, the data sugges t that different diglyceride species produced in response to epidermal growth factor may also induce different biological effects.