title: NF-kappaB activation by the chemopreventive dithiolethione oltipraz is exerted through stimulation of MEKK3 signaling.
abstract: Chemoprevention by the dithiolethione analogue oltipraz (4-methyl-5-(2-pyrazinyl)-1,2-dithiole-3-thione) may occur through several mechanisms, among them stimulation of detoxication activity. The phase II detoxication enzyme, NAD(P)H:quinone oxidoreductase 1 (NQO1; EC 1.6.99.2) also known as quinone reductase (QR) is well established to undergo transcriptional activation following oltipraz treatment of colon cancer cells in culture. Promoter analysis of the QR gene in oltipraztreated cells reveals the involvement of both the AP-1 and NF-kappaB elements in the response. The emerging role of NF-kappaB in cell survival prompted a fuller analysis of effects of oltipraz on this pathway. Oltipraz treatment of both HCT116 and HT29 cells results in the induction of proteins involved in both pathways of NF-kappaB activation, including p65, IkappaB kinase alpha (IKKalpha), IkappaB kinase beta (IKKbeta), and NF-kappaB-inducing kinase (NIK). IkappaBalpha total protein levels were unchanged, but phosphorylation of the inhibitor was also induced in both lines. Electrophoretic mobility shift assay (EMSA) analysis confirmed induction of protein binding to a consensus NF-kappaB element, and transcriptional activation was further confirmed using a reporter construct. Transcriptional activation of QR was decreased in a dose-dependent manner by dominant-negative NF-kappaB in both cell lines. The molecular mechanism that triggers IKK activation in response to oltipraz was also examined using inhibitory constructs of NIK and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 3 (MEKK3). We found that both MEKK3 and NIK exert effects on IKKalpha/beta activation, but through different pathways. Furthermore, the receptor-interacting protein (RIP) was found to interact strongly with MEKK3 during oltipraz-induced NF-kappaB signaling, implying a role for tumor necrosis factor receptor signaling in the action of oltipraz. These results implicate a novel signaling pathway for the action of oltipraz in QR gene regulation.
authors.affiliation: Division of Hematology-Oncology, School of Medicine and Abramson Cancer Center, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Chu Won
authors.initials: CW
authors.lastname: Nho
authors.forename: Peter J
authors.initials: PJ
authors.lastname: O'Dwyer
mesh_terms: D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D002470:Cell Survival; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation; D005799:Genes, Dominant; D017930:Genes, Reporter; D006801:Humans; D051550:I-kappa B Kinase; D048748:MAP Kinase Kinase Kinase 3; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism*; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D011719:Pyrazines / Q000494:pharmacology*; D034741:RNA, Small Interfering / Q000378:metabolism; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; D015398:Signal Transduction; D013871:Thiones; D013876:Thiophenes; D013997:Time Factors; D015533:Transcriptional Activation; D014162:Transfection; D000097782:NF-kappaB-Inducing Kinase
chemical_list: D016588:Anticarcinogenic Agents; D051550:I-kappa B Kinase; D048748:MAP Kinase Kinase Kinase 3; D020930:MAP Kinase Kinase Kinases; D016660:NAD(P)H Dehydrogenase (Quinone); D016328:NF-kappa B; D017346:Protein Serine-Threonine Kinases; D011506:Proteins; D011719:Pyrazines; D034741:RNA, Small Interfering; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; D013871:Thiones; D013876:Thiophenes; D000097782:NF-kappaB-Inducing Kinase; C026209:oltipraz; C447479:NQO1 protein, human; C506249:RIPK1 protein, human; C496559:CHUK protein, human; C496560:IKBKB protein, human; C496565:IKBKE protein, human; C482625:MAP3K3 protein, human