INVOLVEMENT OF THE RAS EXTRACELLULAR SIGNAL-REGULATED KINASE SIGNALING PATHWAY IN THE REGULATION OF ERCC-1 MESSENGER-RNA LEVELS BY INSULIN

Expression of DNA repair enzymes, which includes ERCC-1, might be unde r the control of hormonal and growth factor stimulation. In the presen t study it was observed that insulin increased ERCC-1 mRNA levels both in Chinese hamster ovary cells overexpressing human insulin receptors (HIRc cells) and in fully differentiated 3T3-L1 adipocytes, To invest igate the mechanisms underlying the increase in ERCC-1 gene expression in HIRc cells, we used a variety of pharmacological tools known to in hibit distinct signalling pathways. None of these inhibitors affected the amount of ERCC-1 mRNA in unstimulated cells. The pretreatment of c ells with two chemically unrelated phosphatidylinositol 3'-kinase inhi bitors, wortmannin and LY294002, failed to block the doubling of ERCC- 1 mRNA content by insulin. Similarly, inhibition of pp70 S6 kinase by rapamycin had no apparent effects on this insulin response. In contras t, altering the p21(ras)-dependent pathway with either manumycin, an i nhibitor of Ras farnesylation, or PD98059, an inhibitor of the mitogen -activated protein kinase/extracellular signal-regulated protein kinas e (ERK) kinase, suppressed the induction of ERCC-1 mRNA by insulin (P < 0.001), Furthermore inhibition of RNA and protein synthesis negative ly regulated the expression of this insulin-regulated gene (P < 0.005) . These results suggest that insulin enhances ERCC-1 mRNA levels by th e activation of the Ras-ERK-dependent pathway without the involvement of the phosphatidylinositol 3'-kinase/pp70 S6 kinase.