Involvement of the Ras/extracellular signal-regulated kinase signalling pathway in the regulation of ERCC-1 mRNA levels by insulin.
Expression of DNA repair enzymes, which includes ERCC-1, might be under the control of hormonal and growth factor stimulation. In the present 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 investigate the mechanisms underlying the increase in ERCC-1 gene expression in HIRc cells, we used a variety of pharmacological tools known to inhibit distinct signalling pathways. None of these inhibitors affected the amount of ERCC-1 mRNA in unstimulated cells. The pretreatment of cells with two chemically unrelated phosphatidylinositol 3'-kinase inhibitors, 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 contrast, altering the p21(ras)-dependent pathway with either manumycin, an inhibitor of Ras farnesylation, or PD98059, an inhibitor of the mitogen-activated protein kinase/extracellular signal-regulated protein kinase (ERK) kinase, suppressed the induction of ERCC-1 mRNA by insulin (P<0.001). Furthermore inhibition of RNA and protein synthesis negatively regulated the expression of this insulin-regulated gene (P<0.005). These results suggest that insulin enhances ERCC-1 mRNA levels by the activation of the Ras-ERK-dependent pathway without the involvement of the phosphatidylinositol 3'-kinase/pp70 S6 kinase.
Lee-Kwon
W
W
Diabetes Section, Laboratory of Clinical Investigation, National Institute on Aging, National Institutes of Health, 5600 Nathan Shock Drive, Baltimore, MD 21224, USA.
Park
D
D
Bernier
M
M
D017667:Adipocytes / Q000378:metabolism; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D016466:CHO Cells; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D002867:Chromones / Q000494:pharmacology; D006224:Cricetinae; D004268:DNA-Binding Proteins; D004720:Endonucleases; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D009025:Morpholines / Q000494:pharmacology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011090:Polyenes / Q000494:pharmacology; D053284:Polyunsaturated Alkamides; D017368:Protein Prenylation / Q000187:drug effects; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism; D019893:Ribosomal Protein S6 Kinases / Q000037:antagonists & inhibitors; D015398:Signal Transduction; D000077191:Wortmannin; D018631:ras Proteins / Q000378:metabolism*
D000730:Androstadienes; D002867:Chromones; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D005419:Flavonoids; D007328:Insulin; D009025:Morpholines; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011090:Polyenes; D053284:Polyunsaturated Alkamides; D011506:Proteins; D012333:RNA, Messenger; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D011972:Receptor, Insulin; D019893:Ribosomal Protein S6 Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; C071447:ERCC1 protein, human; D004720:Endonucleases; D018631:ras Proteins; C054474:manumycin; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; D000077191:Wortmannin