Nck recruitment to Eph receptor, EphB1/ELK, couples ligand activation to c-Jun kinase.
Eph family receptor tyrosine kinases signal axonal guidance, neuronal bundling, and angiogenesis; yet the signaling systems that couple these receptors to targeting and cell-cell assembly responses are incompletely defined. Functional links to regulators of cytoskeletal structure are anticipated based on receptor mediated cell-cell aggregation and migratory responses. We used two-hybrid interaction cloning to identify EphB1-interactive proteins. Six independent cDNAs encoding the SH2 domain of the adapter protein, Nck, were recovered in a screen of a murine embryonic library. We mapped the EphB1 subdomain that binds Nck and its Drosophila homologue, DOCK, to the juxtamembrane region. Within this subdomain, Tyr594 was required for Nck binding. In P19 embryonal carcinoma cells, activation of EphB1 (ELK) by its ligand, ephrin-B1/Fc, recruited Nck to native receptor complexes and activated c-Jun kinase (JNK/SAPK). Transient overexpression of mutant EphB1 receptors (Y594F) blocked Nck recruitment to EphB1, attenuated downstream JNK activation, and blocked cell attachment responses. These findings identify Nck as an important intermediary linking EphB1 signaling to JNK.
Stein
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Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
Huynh-Do
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Lane
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Cerretti
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DP
Daniel
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TO
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D019556:COS Cells; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002448:Cell Adhesion; D029721:Drosophila Proteins; D004789:Enzyme Activation; D036387:Ephrin-B1; D005353:Fibronectins / Q000378:metabolism; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases; D008024:Ligands; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000378:metabolism; D015513:Oncogene Proteins / Q000378:metabolism*; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D029721:Drosophila Proteins; D036387:Ephrin-B1; D005353:Fibronectins; D008024:Ligands; D008565:Membrane Proteins; C078332:Nck protein; D009419:Nerve Tissue Proteins; D015513:Oncogene Proteins; C099753:dock protein, Drosophila; D014443:Tyrosine; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases