title: Claudin-2 knockdown decreases matrix metalloproteinase-9 activity and cell migration via suppression of nuclear Sp1 in A549 cells.
abstract: OBJECTIVE
  Claudin expression is altered in lung cancer, but the pathophysiological role of claudin is not well understood. We examined the effect of claudin-2 expression on cell migration using human adenocarcinoma A549 cells.
  
  
  METHODS
  The mRNA level was measured by real time polymerase chain reaction. To knockdown claudin-2 expression, we made the cells expressing doxycycline-inducible claudin-2 shRNA vector. The protein level was examined by Western blotting. Cell migration was measured by wound-healing assay. The enzymatic activity of MMP-9 was assessed by gelatin zymography.
  
  
  RESULTS
  In A549 cells, claudin-2 expression was higher than in normal lung tissue. Claudin-2 knockdown did not affect the expression of other junctional proteins including claudin-1, occludin and E-cadherin. Claudin-2 knockdown decreased cell migration concomitant with a decrease in the mRNA level and enzymatic activity of MMP-9. The expression level of Sp1 in the nuclei was decreased by claudin-2 knockdown. In contrast, the expression levels of c-Fos, c-Jun and NF-kB p65 in the nuclei were not changed by claudin-2 knockdown. The knockdown of Sp1 expression by siRNA decreased cell migration, and the mRNA expression, enzymatic activity, and promoter activity of MMP-9.
  
  
  CONCLUSIONS
  Claudin-2 may increase the mRNA level and enzymatic activity of MMP-9 mediated by the elevation of nuclear distribution of Sp1, resulting in the up-regulation of A549 cell migration.
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: Ayumi
authors.initials: A
authors.lastname: Takiguchi
authors.forename: Kosuke
authors.initials: K
authors.lastname: Atomi
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Yamazaki
authors.forename: Junko
authors.initials: J
authors.lastname: Sugatani
mesh_terms: D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D002467:Cell Nucleus / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D057167:Claudins; D003593:Cytoplasm / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D055785:Gene Knockdown Techniques; D006801:Humans; D008168:Lung / Q000201:enzymology / Q000378:metabolism; D008175:Lung Neoplasms / Q000201:enzymology / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D018613:Microscopy, Confocal; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016329:Sp1 Transcription Factor / Q000037:antagonists & inhibitors* / Q000235:genetics; D014162:Transfection
chemical_list: C491875:CLDN2 protein, human; D057167:Claudins; D008565:Membrane Proteins; D016329:Sp1 Transcription Factor; D020780:Matrix Metalloproteinase 9