OXIDIZED LOW-DENSITY LIPOPROTEINS STIMULATE GALACTOSYLTRANSFERASE ACTIVITY, RAS ACTIVATION, P(44) MITOGEN-ACTIVATED PROTEIN-KINASE AND C-FOS EXPRESSION IN AORTIC SMOOTH-MUSCLE CELLS

Previously, our laboratory has shown that oxidized low density lipopro teins (Ox-LDL) can exert a concentration-dependent stimulation in the proliferation of aortic smooth muscle cells, ''a hallmark in the patho genesis of atherosclerosis'' (Chatterjee,S. (1992) Mel. Cell, Biochem. , 111, 143-147), Here we report a novel aspect of Ox-LDL-mediated sign al transduction, We demonstrate that in aortic smooth muscle cells, Ox -LDL stimulates the activity of a UDP-galactose:glucosylceramide beta 1-->4 galactosyltransferase (GalT-2) and phosphorylation/activation of p(44) mitogen-activated protein (MAP) kinase (p(44) MAPK). The activi ty of GalT-2 increased about 2-fold within 2.5-5 min of incubation of cells with Ox-LDL (10 mu g/ml), After 5 min of incubation of cells wit h Ox-LDL, but not LDL, there was a 2-fold increase in the activity of p(44) MAPK, Phosphoamino acid analysis employing thin layer chromatogr aphy revealed that the tyrosine and threonine moieties of,44 MAPK was phosphorylated by Ox-LDL. D-1-Phenyl-2-decanoylamino-3-morpholino-1-pr opanol (D-PDMP; a potent inhibitor of GalT-2) impaired the Ox-LDL medi ated induction of p(44) MAPK activity and the phosphorylation of tyros ine and threonine residues in p(44) MAPK, This phenomenon was bypassed by the simultaneous addition of lactosylceramide. The upstream and do wnstream parameters in MAP kinase signaling pathways were investigated next, We found that Ox-LDL stimulated (9-fold) the loading of GTP on Pas, Interestingly, Ox-LDL specifically induced c-fos mRNA expression (6.5-fold) in these cells, as compared to the control, Thus, one of th e biochemical mechanisms in Ox-LDL mediated induction in the prolifera tion in aortic smooth muscle cells may involve GalT-2 activation, lact osylceramide production, Pas GTP loading, activation of the kinase cas cade, and c-fos expression.