title: P130Cas attenuates epidermal growth factor (EGF) receptor internalization by modulating EGF-triggered dynamin phosphorylation.
abstract: BACKGROUND
  Endocytosis controls localization-specific signal transduction via epidermal growth factor receptor (EGFR), as well as downregulation of that receptor. Extracellular matrix (ECM)-integrin coupling induces formation of macromolecular complexes that include EGFR, integrin, Src kinase and p130Cas, resulting in EGFR activation. In addition, cell adhesion to ECM increases EGFR localization at the cell surface and reduces EGFR internalization. The molecular mechanisms involved are not yet well understood.
  
  
  RESULTS
  We investigated the molecular mechanism by which p130Cas affects the endocytic regulation of EGFR. Biochemical quantification revealed that cell adhesion to fibronectin (FN) increases total EGFR levels and its phosphorylation, and that p130Cas is required for this process. Measurements of Texas Red-labeled EGF uptake and cell surface EGFR revealed that p130Cas overexpression reduces EGF-induced EGFR internalization, while p130Cas depletion enhances it. In addition, both FN-mediated cell adhesion and p130Cas overexpression reduce EGF-stimulated dynamin phosphorylation, which is necessary for EGF-induced EGFR internalization. Coimmunoprecipitation and GST pull-down assays confirmed the interaction between p130Cas and dynamin. Moreover, a SH3-domain-deleted form of p130Cas, which shows diminished binding to dynamin, inhibits dynamin phosphorylation and EGF uptake less effectively than wild-type p130Cas.
  
  
  CONCLUSIONS
  Our results show that p130Cas plays an inhibitory role in EGFR internalization via its interaction with dynamin. Given that the EGFR internalization process determines signaling density and specificity in the EGFR pathway, these findings suggest that the interaction between p130Cas and dynamin may regulate EGFR trafficking and signaling in the same manner as other endocytic regulatory proteins related to EGFR endocytosis.
authors.affiliation: Department of Life Science, Bio Imaging and Cell Dynamics Research Center, Gwangju Institute of Science and Technology, Gwangju, Korea.
authors.forename: Yong Seok
authors.initials: YS
authors.lastname: Kang
authors.forename: Wook
authors.initials: W
authors.lastname: Kim
authors.forename: Yun Hyun
authors.initials: YH
authors.lastname: Huh
authors.forename: Jeomil
authors.initials: J
authors.lastname: Bae
authors.forename: Jin Soo
authors.initials: JS
authors.lastname: Kim
authors.forename: Woo Keun
authors.initials: WK
authors.lastname: Song
mesh_terms: D000818:Animals; D015153:Blotting, Western; D050719:Crk-Associated Substrate Protein / Q000502:physiology*; D034281:Dynamins / Q000378:metabolism*; D004705:Endocytosis; D004815:Epidermal Growth Factor / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D010766:Phosphorylation
chemical_list: D050719:Crk-Associated Substrate Protein; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D034281:Dynamins