TY - JOUR
T1 - Phosphorylation of c-Crk II on the negative regulatory Tyr222 mediates nerve growth factor-induced cell spreading and morphogenesis
AU - Escalante, M.
AU - Courtney, J.
AU - Wai Gong Chin, Gong Chin
AU - Teng, K. K.
AU - Kim, J. I.
AU - Fajardo, J. E.
AU - Mayer, B. J.
AU - Hempstead, B. L.
AU - Birge, R. B.
PY - 2000/8/11
Y1 - 2000/8/11
N2 - 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.
AB - 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.
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U2 - 10.1074/jbc.M000711200
DO - 10.1074/jbc.M000711200
M3 - Article
C2 - 10825157
AN - SCOPUS:0034637444
SN - 0021-9258
VL - 275
SP - 24787
EP - 24797
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 32
ER -