title: Thioredoxin inhibits MPK38-induced ASK1, TGF-β, and p53 function in a phosphorylation-dependent manner.
abstract: Murine protein serine-threonine kinase 38 (MPK38) is a member of the AMP-activated protein kinase-related serine/threonine kinase family. The factors that regulate MPK38 activity and function are not yet elucidated. Here, thioredoxin (Trx) was shown to be a negative regulator of MPK38. The redox-dependent association of MPK38 and Trx was mediated through the C-terminal domain of MPK38. Single and double amino acid substitution mutagenesis of MPK38 (C286S, C339S, C377S, and C339S/C377S) and Trx (C32S, C35S, and C32S/C35S) demonstrated that Cys(339) and Cys(377) of MPK38 and Cys(32) and Cys(35) of Trx are required for MPK38-Trx complex formation. MPK38 directly interacted with and phosphorylated Trx at Thr(76). Expression of wild-type Trx, but not the Trx mutants C32S/C35S and T76A, inhibited MPK38-induced ASK1, TGF-β, and p53 function by destabilizing MPK38. The E3 ubiquitin-protein ligase Mdm2 played a critical role in the regulation of MPK38 stability by Trx. Treatment of cells with 1-chloro-2,4-dinitrobenzene, a specific inhibitor of Trx reductase, decreased MPK38-Trx complex formation and subsequently increased MPK38 stability and activity, indicating that Trx negatively regulates MPK38 activity in vivo. Finally, we used ASK1-, Smad3-, and p53-null mouse embryonic fibroblasts to demonstrate that ASK1, Smad3, and p53 play important roles in the activity and function of MPK38, suggesting a functional link between MPK38 and ASK1, TGF-β, and p53 signaling pathways. These results indicate that Trx functions as a physiological inhibitor of MPK38, which plays an important role in inducing ASK1-, TGF-β-, and p53-mediated activity.
authors.affiliation: Department of Biochemistry, School of Life Sciences, Chungbuk National University, Cheongju 361-763, Korea.
authors.forename: Ravi
authors.initials: R
authors.lastname: Manoharan
authors.forename: Hyun-A
authors.initials: HA
authors.lastname: Seong
authors.forename: Hyunjung
authors.initials: H
authors.lastname: Ha
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D006801:Humans; D048848:MAP Kinase Kinase Kinase 5 / Q000378:metabolism*; D051379:Mice; D016296:Mutagenesis; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000378:metabolism; D013879:Thioredoxins / Q000235:genetics* / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
chemical_list: D016212:Transforming Growth Factor beta; D016159:Tumor Suppressor Protein p53; D013879:Thioredoxins; C108524:Melk protein, mouse; D017346:Protein Serine-Threonine Kinases; D048848:MAP Kinase Kinase Kinase 5
keywords: ASK1; Free radicals; MPK38/MELK; TGF-β; Trx; p53