Phosphorylation of c-Crk II on the negative regulatory Tyr222 mediates nerve growth factor-induced cell spreading and morphogenesis

M. Escalante, J. Courtney, Gong Chin Wai Gong Chin, K. K. Teng, J. I. Kim, J. E. Fajardo, B. J. Mayer, B. L. Hempstead, R. B. Birge

Research output: Contribution to journalArticle

44 Scopus citations

Abstract

The Crk family of adaptor proteins participate in diverse signaling pathways that regulate growth factor induced proliferation, anchorage-dependent DNA synthesis, and cytoskeletal reorganization, important for cell adhesion and motility. Using kidney epithelial 293T cells for transient co-transfection studies and the nerve growth factor (NGF)-responsive PC12 cell line as a model system for neuronal morphogenesis, we demonstrate that the non-receptor tyrosine kinase c-Abl is an intermediary for NGF-inducible c-Crk II phosphorylation on the negative regulatory Tyr222. Transient expression of a c-Crk II Tyr222 point mutant (c-Crk Y222F) in 293T cells induces hyperphosphorylation of paxillin on Tyr31 and enhances complex formation between c-Crk Y222F and paxillin as well as c-Crk Y222F and c-Abl, suggesting that c-Crk II Tyr222 phosphorylation induces both the dissociation of the Crk SH2 domain from paxillin and the Crk SH3do domain from c-Abl. Interestingly, examination of the early kinetics of NGF stimulation in PC12 cells showed that c-Crk II Tyr222 phosphorylation preceded paxillin Tyr31 phosphorylation, followed by a transient initial dissociation of the c-Crk II paxillin complex. PC12 cells overexpressing c-Crk Y222F manifested a defect in cellular adhesion and neuritogenesis that led to detachment of cells from the extracellular matrix, thus demonstrating the biological significance of c-Crk II tyrosine phosphorylation in NGF-dependent morphogenesis. Whereas previous studies have shown that Crk SH2 binding to paxillin is critical for cell adhesion and migration, our data show that the phosphorylation cycle of c-Crk II determines its dynamic interaction with paxillin, thereby regulating turnover of multiprotein complexes, a critical aspect of cytoskeletal plasticity and actin dynamics.

Original languageEnglish (US)
Pages (from-to)24787-24797
Number of pages11
JournalJournal of Biological Chemistry
Volume275
Issue number32
DOIs
StatePublished - Aug 11 2000
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Escalante, M., Courtney, J., Wai Gong Chin, G. C., Teng, K. K., Kim, J. I., Fajardo, J. E., Mayer, B. J., Hempstead, B. L., & Birge, R. B. (2000). Phosphorylation of c-Crk II on the negative regulatory Tyr222 mediates nerve growth factor-induced cell spreading and morphogenesis. Journal of Biological Chemistry, 275(32), 24787-24797. https://doi.org/10.1074/jbc.M000711200