title: Epidermal growth factor increases clathrin-dependent endocytosis and degradation of claudin-2 protein in MDCK II cells.
abstract: Epidermal growth factor (EGF) decreases the mRNA and protein levels of claudin-2 (CLDN2) in Madin-Darby canine kidney (MDCK) II cells. Here we examined whether EGF affects the stability and intracellular distribution of CLDN2 protein. EGF decreased surface and total levels of CLDN2, which was inhibited by U0126, a MEK inhibitor. CLDN2 was co-localized at the tight junctions (TJs) with ZO-1, a scaffolding protein. The fluorescence signal for CLDN2 disappeared on treatment with EGF, which was inhibited by U0126. EGF accelerated the decrease in CLDN2 in the presence of cycloheximide, a translation inhibitor, indicating that EGF reduces the stability of the protein. Chloroquine, a lysosomal protease inhibitor, blocked the EGF-induced decrease in CLDN2 protein and caused the co-localization of CLDN2 with Lamp-1, a marker of lysosome. Monodancylcadaverine, an inhibitor of endocytosis, and clathrin siRNA blocked the EGF-induced decrease in CLDN2 and the translocation of CLDN2 from the TJs to the lysosome. EGF increased the association of CLDN2 with clathrin and adaptin α which was inhibited by U0126. These results suggest that EGF accelerates clathrin-dependent endocytosis and lysosomal degradation of CLDN2 protein mediated by the activation of a MEK/ERK pathway.
authors.affiliation: Department of Pharmaco-Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka, Japan. ikari@u-shizuoka-ken.ac.jp
authors.forename: Akira
authors.initials: A
authors.lastname: Ikari
authors.forename: Ayumi
authors.initials: A
authors.lastname: Takiguchi
authors.forename: Kosuke
authors.initials: K
authors.lastname: Atomi
authors.forename: Junko
authors.initials: J
authors.lastname: Sugatani
mesh_terms: D000818:Animals; D002103:Cadaverine / Q000031:analogs & derivatives / Q000494:pharmacology; D002460:Cell Line; D002738:Chloroquine / Q000494:pharmacology; D002966:Clathrin / Q000378:metabolism*; D004285:Dogs; D004705:Endocytosis / Q000187:drug effects*; D004789:Enzyme Activation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D052118:Lysosomal-Associated Membrane Protein 1 / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects*; D021381:Protein Transport / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D019108:Tight Junctions / Q000187:drug effects / Q000378:metabolism
chemical_list: D002966:Clathrin; D052118:Lysosomal-Associated Membrane Protein 1; D008565:Membrane Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D004815:Epidermal Growth Factor; D002738:Chloroquine; D048049:Extracellular Signal-Regulated MAP Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; C008542:monodansylcadaverine; D002103:Cadaverine