Substrate specificity for normal but not mutationally activated variants of src family kinases.
Although structural features and expression patterns of the src family of tyrosine kinases have been extensively analyzed, there are no direct comparative studies of the putative protein substrates that are tyrosine-phosphorylated by the normal cellular versions of these enzymes. In this report, we have expressed normal and enzymatically activated versions of the fyn, fgr, and src translational products by transfection of appropriate cDNAs into mouse fibroblasts. Because the same parental cell line was used for all transfections, each enzyme was expressed in a similar milieu of potential in vivo substrates. After verification of appropriate expression from each transfected cDNA and assessment of relative transforming potency, a series of putative protein substrates was specifically assayed for expression and tyrosine phosphorylation. Our data indicate that the normal src family kinases display some degree of substrate specificity but that specificity is diminished when these enzymes are constitutively activated. In the course of these studies, tyrosine-phosphorylated proteins were noted to coimmunoprecipitate with some of these putative in vivo substrates. Some of these coimmunoprecipitating proteins have been reported previously, whereas others, such as the presence of p59fyn in anti-p80/85 immunoprecipitates, are heretofore undescribed.
Sartor
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Clinical Pharmacology Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892.
Robbins
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KC
D016475:3T3 Cells; D000818:Animals; D000081247:CSK Tyrosine-Protein Kinase; D004789:Enzyme Activation; D051379:Mice; D009154:Mutation; D010766:Phosphorylation; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D051076:Proto-Oncogene Proteins c-fyn; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism; D013379:Substrate Specificity; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases
D011518:Proto-Oncogene Proteins; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; C494960:Fyn protein, mouse; D051076:Proto-Oncogene Proteins c-fyn; D016392:Proto-Oncogene Proteins pp60(c-src); C053102:proto-oncogene proteins c-fgr; D019061:src-Family Kinases