title: Claudin-4 controls the receptor tyrosine kinase EphA2 pro-oncogenic switch through β-catenin.
abstract: BACKGROUND
  The EphA2 receptor, which is expressed in many types of cancer, is activated by two different mechanisms. Activation by engagement with one of its ephrin ligands is anti-oncogenic whereas phosphorylation of S897 by AKT increases migration, invasion and metastasis. Down-regulation of claudin-4 (CLDN4) produces a loss of E-cadherin and increased β-catenin signaling and a phenotype similar to that produced by oncogenic activation of EphA2, suggesting that CLDN4 may serve to restrain the pro-oncogenic signaling of EphA2.
  
  
  RESULTS
  We found that constitutive knockdown of CLDN4 was associated with a 4.5-fold increase in EphA2 mRNA and a 2.5-fold increase in EphA2 protein which was reversible by re-expression of CLDN4. Knockdown of EphA2 blocked the migratory phenotype induced by loss of CLDN4. Knockdown of CLDN4 resulted in a 5.8-fold increase in pEphA(S897), the oncogenic form of the receptor, as well as partial mislocalization of the excess EphA2 to the interior of the cell. Forced expression of E-cadherin did not reduce total EphA2 or pEphA(S897) whereas re-expression of CLDN4 restored localization and reduced EphA2 and pEphA(S897) even in cells not expressing E-cadherin. Transient siRNA-mediated knockdown of EphA2 and β-catenin, and inhibition of PI3K by LY294002, demonstrated that increased pEphA(S897) in the CLDN4 knockdown cells was attributable to an increase in the level of active dephospho-β-catenin upstream of PI3K and AKT.
  
  
  CONCLUSIONS
  We conclude that CLDN4 serves to restrain pro-oncogenic signaling from EphA2 by limiting the activity of β-catenin and PI3K and preventing phosphorylation of EphA2 on S897 by AKT. This suggests that interventions directed at enhancing the level or functional activity of CLDN4 may be of therapeutic interest.
authors.affiliation: Department of Medicine and the Moores UCSD Cancer Center, University of California, 3855 Health Sciences Drive, La Jolla, San Diego, CA, 92093-0819, USA. xshang@ucsd.edu.
authors.forename: Xiying
authors.initials: X
authors.lastname: Shang
authors.affiliation: Department of Medicine and the Moores UCSD Cancer Center, University of California, 3855 Health Sciences Drive, La Jolla, San Diego, CA, 92093-0819, USA. xlin@ucsd.edu.
authors.forename: Xinjian
authors.initials: X
authors.lastname: Lin
authors.affiliation: Department of Medicine and the Moores UCSD Cancer Center, University of California, 3855 Health Sciences Drive, La Jolla, San Diego, CA, 92093-0819, USA. showell@ucsd.edu.
authors.forename: Stephen B
authors.initials: SB
authors.lastname: Howell
mesh_terms: D045744:Cell Line, Tumor; D002465:Cell Movement; D062506:Claudin-4 / Q000235:genetics / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006801:Humans; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D036104:Receptor, EphA2 / Q000235:genetics / Q000378:metabolism*; D014945:Wound Healing; D051176:beta Catenin / Q000378:metabolism*
chemical_list: D062506:Claudin-4; D034741:RNA, Small Interfering; D051176:beta Catenin; D019869:Phosphatidylinositol 3-Kinases; D036104:Receptor, EphA2; D051057:Proto-Oncogene Proteins c-akt