Paxillin phosphorylation and association with Lck and Pyk2 in anti-CD3- or anti-CD45-stimulated T cells.
Antibodies to either CD3 or CD45 have been shown to induce dramatic changes in cell morphology, increased tyrosine phosphorylation of cellular proteins, and the association of a subset of these proteins with the tyrosine kinase Lck. The current study was initiated to determine the identity of the tyrosine-phosphorylated 70-80 kDa protein that becomes Lck-associated after stimulation with anti-CD45 or anti-CD3. We demonstrate that the cytoskeletal protein paxillin becomes tyrosine-phosphorylated when cells are plated on immobilized antibodies specific for CD45 or CD3. Only tyrosine-phosphorylated paxillin is associated with Lck, suggesting that the association is through the SH2 domain of Lck. Consistent with this we demonstrate that the SH2 domain of Lck binds tyrosine-phosphorylated paxillin. In contrast, the association of paxillin with the FAK-related kinase Pyk2 was found to be constitutive and not altered by the phosphorylation of either protein. Finally, we establish that the phosphorylation of paxillin is dependent on the expression of Lck. Taken together, these results demonstrate that paxillin is physically associated with kinases from two different families in T cells and suggest that paxillin may function as an adaptor protein linking cellular signals with cytoskeletal changes during T cell activation.
Ostergaard
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Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta T6G 2H7, Canada. ostergaard@ualberta.ca
Lou
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Arendt
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Berg
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D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000494:pharmacology; D017252:CD3 Complex / Q000276:immunology*; D002460:Cell Line; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D003599:Cytoskeleton / Q000502:physiology; D051418:Focal Adhesion Kinase 2; D017493:Leukocyte Common Antigens / Q000276:immunology*; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / Q000378:metabolism*; D051379:Mice; D010755:Organophosphates / Q000494:pharmacology; D051419:Paxillin; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000032:analysis; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D013602:T-Lymphocytes, Cytotoxic / Q000502:physiology*; D018909:src Homology Domains / Q000502:physiology
D000906:Antibodies; D017252:CD3 Complex; D003598:Cytoskeletal Proteins; D010755:Organophosphates; D051419:Paxillin; D010750:Phosphoproteins; C496062:Pxn protein, mouse; D019000:Phosphotyrosine; C074782:phenylphosphate; D011505:Protein-Tyrosine Kinases; D051418:Focal Adhesion Kinase 2; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); C496053:Ptk2b protein, mouse; D017493:Leukocyte Common Antigens