LOVASTATIN INHIBITS THE STIMULATION OF MITOGEN-ACTIVATED PROTEIN-KINASE BY INSULIN IN HIRCB FIBROBLASTS

Lovastatin, a cholesterol biosynthesis inhibitor, has recently been sh own to inhibit mitogenesis and tumor growth, We have investigated the effects of lovastatin on the activation of MAP kinase by insulin using as a model HIRcB cells, a rat fibroblast cell line that overexpresses the human insulin receptor, Treatment with lovastatin (1-30 mu M) for 24 h decreased the level of activation of MAP kinase by insulin by as much as 60%, Immunoblotting experiments using a specific anti-MAP kin ase monoclonal antibody demonstrated that the amount of MAP kinase pro tein in the cells was not altered by lovastatin treatment, Likewise, l ovastatin had no apparent effects on the expression of the insulin rec eptor, Treatment with lovastatin (20 mu M) reduced the percentage of f arnesylated Ras by 50%, Immunoprecipitation of tyrosine phosphorylated proteins from HIRcB cell lysates followed by immunodetection of MAP k inase using specific antibodies demonstrated a reduced level of insuli n-induced tyrosine phosphorylation levels of MAP kinase in lovastatin- treated cells, Furthermore, immunodetection of the beta-subunit of the insulin receptor in anti-phosphotyrosine immunoprecipitates revealed that treatment with lovastatin reduced the tyrosine phosphorylation le vels of the receptor, Lysates obtained from cells treated with increas ing concentrations of lovastatin demonstrated a dose-dependent inhibit ion of the insulin-induced tyrosine phosphorylation of the receptor, T reatment with mevalonic acid prevented the effects of lovastatin demon strating that the effects of the drug are a consequence of its inhibit ory effects on the synthesis of steroids, It is concluded that, in add ition to inhibition of Ras farnesylation, lovastatin reduces receptor tyrosine phosphorylation levels which also contributes to the blockade of MAPK activation by the insulin receptor. (C) 1996 Academic Press, Inc.