title: ERK1/2-driven and MKP-mediated inhibition of EGF-induced ERK5 signaling in human proximal tubular cells.
abstract: The MEK1-ERK1/2 signaling pathway has been implicated in the regulation of renal epithelial cell proliferation, epithelial-to-mesenchymal transition and the induction of an invasive cell phenotype. Much less information is available about the MEK5-ERK5 module and its role in renal epithelial cell proliferation and differentiation. In the present study we have investigated the regulation of these two families of extracellular signal-regulated kinases in epidermal growth factor (EGF)-stimulated human kidney-2 (HK-2) cells and a possible interaction between ERK1/2 and ERK5. Here we report that 5 ng/ml EGF led to a strong stimulation of HK-2 cell proliferation, which was largely U0126-sensitive. Both synthetic MEK1/2 inhibitors U0126 and Cl-1040, when used at 10 and 1 microM, respectively, inhibited basal and EGF-induced ERK1/2 phosphorylation but not ERK5 phosphorylation. Long-term inhibition of MEK1/2-ERK1/2 signaling and/or vanadate-sensitive protein phosphatases enhanced and prolonged EGF-induced ERK5 phosphorylation, while transient expression of an adenoviral constitutively active MEK1 (Ad-caMEK1) construct completely blocked EGF-induced ERK5 phosphorylation. Expression of Ad-caMEK1 in HK-2 cells resulted in the upregulation of the dual-specificity phosphatases MKP-3/DUSP6, MKP-1/DUSP1, and DUSP5. The EGF-mediated time-dependent induction of MKP-3, MKP-1 and DUSP5 mRNA levels was U0126-sensitive at a concentration, which blocked EGF-mediated ERK1/2 phosphorylation but not ERK5 phosphorylation. Furthermore, U0126 inhibited EGF-induced MKP-3 and MKP-1 protein expression. Both MKP-3 and MKP-1 co-immunoprecipitated with ERK5 in unstimulated as well as in EGF-stimulated HK-2 cells. These results suggest the existence of an ERK1/2-driven negative feed-back regulation of ERK5 signaling in EGF-stimulated HK-2 cells, which is mediated by MKP-3, DUSP5 and/or MKP-1.
authors.affiliation: Clinical Division of Nephrology, Department of Internal Medicine, Medical University of Innsbruck, Innsbruck, Austria.
authors.forename: Rita
authors.initials: R
authors.lastname: Sarközi
authors.forename: Bradley
authors.initials: B
authors.lastname: Miller
authors.forename: Verena
authors.initials: V
authors.lastname: Pollack
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Feifel
authors.forename: Gert
authors.initials: G
authors.lastname: Mayer
authors.forename: Andrey
authors.initials: A
authors.lastname: Sorokin
authors.forename: Herbert
authors.initials: H
authors.lastname: Schramek
mesh_terms: D001549:Benzamides / Q000494:pharmacology; D002070:Butadienes / Q000494:pharmacology; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D054638:Dual Specificity Phosphatase 1; D054642:Dual Specificity Phosphatase 6; D054637:Dual-Specificity Phosphatases; D004789:Enzyme Activation / Q000187:drug effects; D004790:Enzyme Induction / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D048369:MAP Kinase Kinase 1 / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D048054:Mitogen-Activated Protein Kinase 7 / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D054645:Protein Phosphatase 1; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013997:Time Factors; D014638:Vanadates / Q000494:pharmacology
chemical_list: C120227:2-(2-chloro-4-iodophenylamino)-N-cyclopropylmethoxy-3,4-difluorobenzamide; D001549:Benzamides; D002070:Butadienes; D018797:Cell Cycle Proteins; D017874:Immediate-Early Proteins; D007527:Isoenzymes; D009570:Nitriles; D012333:RNA, Messenger; C113580:U 0126; D014638:Vanadates; D004815:Epidermal Growth Factor; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048054:Mitogen-Activated Protein Kinase 7; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; C517019:DUSP1 protein, human; C442320:DUSP5 protein, human; C517035:DUSP6 protein, human; D054638:Dual Specificity Phosphatase 1; D054642:Dual Specificity Phosphatase 6; D054637:Dual-Specificity Phosphatases; D017027:Protein Tyrosine Phosphatases