title: F-box protein complex FBXL19 regulates TGFβ1-induced E-cadherin down-regulation by mediating Rac3 ubiquitination and degradation.
abstract: BACKGROUND
  Rac3 is a small GTPase multifunctional protein that regulates cell adhesion, migration, and differentiation. It has been considered as an oncogene in breast cancer; however, its role in esophageal cancer and the regulation of its stability have not been studied. F-box proteins are major subunits within the Skp1-Cullin-1-F-box (SCF) E3 ubiquitin ligases that recognize particular substrates for ubiquitination and proteasomal degradation. Recently, we have shown that SCFFBXL19 targets Rac1 and RhoA, thus regulating Rac1 and RhoA ubiquitination and degradation. Here, we demonstrate the role of FBXL19 in the regulation of Rac3 site-specific ubiquitination and stability. Expression of TGFβ1 is associated with poor prognosis of esophageal cancer. TGFβ1 reduces tumor suppressor, E-cadherin, expression in various epithelial-derived cancers. Here we investigate the role of FBXL19-mediated Rac3 degradation in TGFβ1-induced E-cadherin down-regulation in esophageal cancer cells.
  
  
  METHODS
  FBXL19-regulated endogenous and over-expressed Rac3 stability were determined by immunoblotting and co-immunoprecipitation. Esophageal cancer cells (OE19 and OE33) were used to investigate TGFβ1-induced E-cadherin down-regulation by Immunoblotting and Immunostaining.
  
  
  RESULTS
  Overexpression of FBXL19 decreased endogenous and over-expressed Rac3 expression by interacting and polyubiquitinating Rac3, while down-regulation of FBXL19 suppressed Rac3 degradation. Lysine166 within Rac3 was identified as an ubiquitination acceptor site. The FBXL19 variant with truncation at the N-terminus resulted in an increase in Rac3 degradation; however, the FBXL19 variant with truncation at the C-terminus lost its ability to interact with Rac3 and ubiquitinate Rac3 protein. Further, we found that Rac3 plays a critical role in TGFβ1-induced E-cadherin down-regulation in esophageal cancer cells. Over-expression of FBXL19 attenuated TGFβ1-induced E-cadherin down-regulation and esophageal cancer cells elongation phenotype.
  
  
  CONCLUSIONS
  Collectively these data unveil that FBXL19 functions as an antagonist of Rac3 by regulating its stability and regulates the TGFβ1-induced E-cadherin down-regulation. This study will provide a new potential therapeutic strategy to regulate TGFβ1 signaling, thus suppressing esophageal tumorigenesis.
authors.forename: Su
authors.initials: S
authors.lastname: Dong
authors.forename: Jing
authors.initials: J
authors.lastname: Zhao
authors.forename: Jianxin
authors.initials: J
authors.lastname: Wei
authors.forename: Rachel K
authors.initials: RK
authors.lastname: Bowser
authors.forename: Andrew
authors.initials: A
authors.lastname: Khoo
authors.forename: Zhonghui
authors.initials: Z
authors.lastname: Liu
authors.forename: James D
authors.initials: JD
authors.lastname: Luketich
authors.forename: Arjun
authors.initials: A
authors.lastname: Pennathur
authors.forename: Haichun
authors.initials: H
authors.lastname: Ma
authors.affiliation: Department of Medicine and the Acute Lung Injury Center of Excellence, University of Pittsburgh, 3459 Fifth Avenue, NW 628MUH, Pittsburgh 15213, PA, USA. zhaoy3@upmc.edu.
authors.forename: Yutong
authors.initials: Y
authors.lastname: Zhao
mesh_terms: D015820:Cadherins / Q000096:biosynthesis* / Q000235:genetics; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D004938:Esophageal Neoplasms / Q000235:genetics* / Q000473:pathology; D044783:F-Box Proteins / Q000235:genetics* / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D059748:Proteolysis; D053773:Transforming Growth Factor beta1 / Q000378:metabolism*; D054875:Ubiquitination; D020744:rac GTP-Binding Proteins / Q000235:genetics
chemical_list: D015820:Cadherins; D004268:DNA-Binding Proteins; D044783:F-Box Proteins; C499971:FBXL19 protein, human; C411563:RAC3 protein, human; C508960:TGFB1 protein, human; D053773:Transforming Growth Factor beta1; D020744:rac GTP-Binding Proteins