Specific association of the beta isoform of the p85 subunit of phosphatidylinositol-3 kinase with the proto-oncogene c-cbl.
Phosphatidylinositol-3 kinase (PI-3 kinase) has been implicated in cellular events such as mitogenic signaling, actin organization, and receptor sorting. The p85 subunit of PI-3 kinase contains multiple domains capable of protein-protein interactions that may contribute to mediate the multiple physiological functions of this enzyme. Here, we demonstrate that antibodies raised against the p85 subunit of PI-3 kinase immunoprecipitate a single tyrosine-phosphorylated protein of 120 kDa (pp120) from lysates of activated Jurkat T cells and A20 B cells. This protein is the only significant phosphotyrosine-containing protein in p85 immunoprecipitates from these cells, and it cannot be detected in immunoprecipitates of other signaling proteins such as PLC gamma. Furthermore, antibodies specific for the beta isoform of p85 but not antibodies specific for the alpha isoform immunoprecipitate this tyrosine-phosphorylated protein. pp120 completely comigrates with the proto-oncogene c-cbl, which is a 120 kDa protein product abundant in lymphoid cells. Furthermore, immunoblots of p85 immunoprecipitates using antibodies raised against c-cbl detect a band at exactly the position of pp120. In addition, p85 can be detected in immunoblots of c-cbl immunoprecipitates. Thus, pp120 appears to correspond to c-cbl. A direct association between c-cbl and p85 can be observed in vitro using a fusion protein comprising the Src homology 2 (SH2) domains of p85, and this binding is abolished by phenyl phosphate, suggesting that the interaction is mediated through phosphotyrosine-SH2 domain interactions. Thus, these results show important functional differences between the alpha and beta isoforms of p85 in vivo and point to c-cbl as a potentially important mediator of some of the functions of PI-3 kinase in intact cells.
Hartley
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D
Program in Molecular Medicine, University of Massachusetts Medical School, Worcester 01655, USA.
Meisner
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H
Corvera
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S
D000818:Animals; D001402:B-Lymphocytes / Q000201:enzymology / Q000378:metabolism; D002478:Cells, Cultured; D006801:Humans; D015151:Immunoblotting; D007527:Isoenzymes / Q000378:metabolism*; D008213:Lymphocyte Activation / Q000502:physiology; D008214:Lymphocytes / Q000201:enzymology / Q000378:metabolism*; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D050721:Proto-Oncogene Proteins c-cbl; D011519:Proto-Oncogenes; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000201:enzymology / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases
D007527:Isoenzymes; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins; D050721:Proto-Oncogene Proteins c-cbl; D014443:Tyrosine; D044767:Ubiquitin-Protein Ligases; C000715470:MAS1 protein, human; C067419:CBL protein, human; C493849:Cbl protein, mouse