title: Insulin induces a transcriptional activation of epiregulin, HB-EGF and amphiregulin, by a PI3K-dependent mechanism: identification of a specific insulin-responsive promoter element.
abstract: Previously we have shown that insulin-stimulation of RT4 bladder cancer cells leads to increased proliferation, which require HER1 activation, and is accompanied by increased mRNA expression of the EGF-ligands heparin-binding EGF-like growth factor (HB-EGF), amphiregulin (AR), and epiregulin (EPI) [D. Ornskov, E. Nexo, B.S. Sorensen, Insulin-induced proliferation of bladder cancer cells is mediated through activation of the epidermal growth factor system, FEBS J. 273 (2006) 5479-5489]. In the present paper, we have investigated the molecular mechanism leading to this insulin-induced expression. We monitored the decay of mRNA after inhibiting transcription with Actinomycin D and demonstrated that the insulin-mediated increase was not caused by enhanced mRNA stability. In untreated cells, HB-EGF mRNA was the least stable, whereas AR and EPI mRNA decayed with slower kinetics. However, promoter analysis of HB-EGF and EPI demonstrated that insulin stimulated transcription. Studies on the EPI promoter identified the insulin-responsive element to be located in the region -564 to -365bp. This region contains potential binding sites for the transcription factors SP1, AP1, and NF-kappaB. Interestingly, all three transcription factors can be activated by PI3K. We demonstrate that the insulin-induced expression of HB-EGF, AR, and EPI mRNA is completely prevented by the specific PI3K inhibitor Wortmannin, suggesting an involvement of the PI3K.
authors.affiliation: Department of Clinical Biochemistry, Aarhus Sygehus University Hospital in Aarhus, Denmark.
authors.forename: Dorthe
authors.initials: D
authors.lastname: Ornskov
authors.forename: Ebba
authors.initials: E
authors.lastname: Nexo
authors.forename: Boe S
authors.initials: BS
authors.lastname: Sorensen
mesh_terms: D066258:Amphiregulin; D000730:Androstadienes / Q000494:pharmacology; D066255:EGF Family of Proteins; D004815:Epidermal Growth Factor / Q000378:metabolism*; D066260:Epiregulin; D006023:Glycoproteins / Q000378:metabolism*; D066257:Heparin-binding EGF-like Growth Factor; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D001749:Urinary Bladder Neoplasms / Q000378:metabolism; D000077191:Wortmannin
chemical_list: D066258:Amphiregulin; D000730:Androstadienes; D066255:EGF Family of Proteins; D004815:Epidermal Growth Factor; D066260:Epiregulin; D006023:Glycoproteins; D066257:Heparin-binding EGF-like Growth Factor; D007328:Insulin; D036341:Intercellular Signaling Peptides and Proteins; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D012333:RNA, Messenger; D000077191:Wortmannin; C587848:AREG protein, human; C587880:EREG protein, human; C587883:HBEGF protein, human