Proline-rich tyrosine kinase-2 activation by beta 1 integrin fibronectin receptor cross-linking and association with paxillin in human natural killer cells.
Recent evidence indicates that integrin ligation results in activation of focal adhesion kinase (pp125FAK), the prototype of a new subfamily of nonreceptor protein tyrosine kinase (PTK), including FAKB and the proline-rich tyrosine kinase 2 (PYK-2), also termed cell adhesion kinase-beta or related adhesion focal tyrosine kinase. We have previously shown that cross-linking of alpha 4 beta 1 and alpha 5 beta 1 fibronectin receptors on human NK cells stimulates tyrosine phosphorylation of two proteins migrating at 105 and 115 kDa. Here we report that cross-linking of beta 1 integrins on human NK cells stimulates tyrosine phosphorylation and PTK activity of PYK-2. PYK-2 tyrosine phosphorylation was maximal at 1 min and started to decline 20 min after stimulation. Engagement of alpha 4 beta 1 and alpha 5 beta 1 either with specific mAbs or after cell adhesion to fibronectin or its 120- and 40-kDa fragments also triggered PYK-2 tyrosine phosphorylation. Stimulation of PYK-2 tyrosine phosphorylation was inhibited by the tyrosine kinase inhibitor herbimycin A, but not by EGTA, indicating that PYK-2 tyrosine phosphorylation is PTK, but not calcium, dependent. We also demonstrate that PYK-2 is constitutively associated with paxillin, which undergoes tyrosine phosphorylation with the same kinetics of PYK-2 upon beta 1 integrin ligation.
Gismondi
A
A
Department of Experimental Medicine and Pathology, University of Rome La Sapienza, Italy.
Bisogno
L
L
Mainiero
F
F
Palmieri
G
G
Piccoli
M
M
Frati
L
L
Santoni
A
A
D000818:Animals; D000936:Antigen-Antibody Complex / Q000378:metabolism; D002118:Calcium / Q000502:physiology; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D051418:Focal Adhesion Kinase 2; D006801:Humans; D039041:Integrin alpha4beta1; D016023:Integrins / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D019169:Jurkat Cells; D007694:Killer Cells, Natural / Q000201:enzymology* / Q000378:metabolism; D016716:PC12 Cells; D051419:Paxillin; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000378:metabolism* / Q000502:physiology; D051381:Rats; D017469:Receptors, Fibronectin / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D016201:Receptors, Lymphocyte Homing / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D014443:Tyrosine / Q000378:metabolism
D000936:Antigen-Antibody Complex; D003598:Cytoskeletal Proteins; D039041:Integrin alpha4beta1; D016023:Integrins; C496061:PXN protein, human; D051419:Paxillin; D010750:Phosphoproteins; C496063:Pxn protein, rat; D017469:Receptors, Fibronectin; D016201:Receptors, Lymphocyte Homing; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D051418:Focal Adhesion Kinase 2; C496054:Ptk2b protein, rat; D002118:Calcium