title: Increase in claudin-2 expression by an EGFR/MEK/ERK/c-Fos pathway in lung adenocarcinoma A549 cells.
abstract: In human adenocarcinoma, claudin-2 expression is higher than that in normal lung tissue, but the regulatory mechanism of its expression has not been clarified. In human adenocarcinoma A549 cells, claudin-2 level time-dependently increased under the control conditions. In contrast, claudin-1 expression remained constant for 24h. The concentration of epidermal growth factor (EGF) in medium time-dependently increased, which was inhibited by matrix metalloproteinase (MMP) inhibitor II, an inhibitor of MMP-1, 3, 7, and 9. MMP inhibitor II decreased claudin-2 and phosphorylated ERK1/2 (p-ERK1/2) levels, which were recovered by EGF. Both claudin-2 and p-ERK1/2 levels were decreased by EGF neutralizing antibody, EGF receptor (EGFR) siRNA, AG1478, an inhibitor of EGFR, U0126, an inhibitor of MEK, and the exogenous expression of dominant negative-MEK. These results suggest that EGF is secreted from A549 cells by MMP and increases claudin-2 expression mediated via the activation of an EGFR/MEK/ERK pathway. The inhibition of the signaling pathway decreased phosphorylated c-Fos and nuclear c-Fos levels. The introduction of c-Fos siRNA decreased claudin-2 level without affecting claudin-1. The promoter activity of human claudin-2 was decreased by AG1478 and U0126. Furthermore, the activity was decreased by the deletion or mutation of the AP-1 binding site of claudin-2 promoter. Chromatin immunoprecipitation and avidin-biotin conjugated DNA assays showed that c-Fos binds to the AP-1 binding site. We suggest that a secreted EGF up-regulates the transcriptional activity of claudin-2 mediated by the activation of an EGFR/MEK/ERK/c-Fos pathway in A549 cells.
authors.affiliation: Department of Pharmaco-Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526, Japan. ikari@u-shizuoka-ken.ac.jp
authors.forename: Akira
authors.initials: A
authors.lastname: Ikari
authors.forename: Tomonari
authors.initials: T
authors.lastname: Sato
authors.forename: Ryo
authors.initials: R
authors.lastname: Watanabe
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Yamazaki
authors.forename: Junko
authors.initials: J
authors.lastname: Sugatani
mesh_terms: D000230:Adenocarcinoma / Q000201:enzymology* / Q000235:genetics*; D000077192:Adenocarcinoma of Lung; D001665:Binding Sites; D002070:Butadienes / Q000494:pharmacology; D045744:Cell Line, Tumor; D057167:Claudins / Q000235:genetics* / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D008175:Lung Neoplasms / Q000201:enzymology* / Q000235:genetics*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008297:Male; D009570:Nitriles / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics; D011480:Protease Inhibitors / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism*; D011799:Quinazolines; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000378:metabolism; D020032:Tyrphostins
chemical_list: D002070:Butadienes; C491875:CLDN2 protein, human; D057167:Claudins; D009570:Nitriles; D011480:Protease Inhibitors; D016760:Proto-Oncogene Proteins c-fos; D011799:Quinazolines; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D018808:Transcription Factor AP-1; D020032:Tyrphostins; C113580:U 0126; C101044:RTKI cpd; D004815:Epidermal Growth Factor; C512478:EGFR protein, human; D066246:ErbB Receptors