The SH2/SH3 domain-containing protein Nck is recognized by certain anti-phospholipase C-gamma 1 monoclonal antibodies, and its phosphorylation on tyrosine is stimulated by platelet-derived growth factor and epidermal growth factor treatment.
In the course of our investigation of phospholipase C (PLC)-gamma 1 phosphorylation by using a set of anti-PLC-gamma 1 monoclonal antibodies (P.-G. Suh, S. H. Ryu, W. C. Choi, K.-Y. Lee, and S. G. Rhee, J. Biol. Chem. 263:14497-14504, 1988), we found that some of these antibodies directly recognize a 47-kDa protein. We show here that this 47-kDa protein is identical to the SH2/SH3-containing protein Nck (J. M. Lehmann, G. Riethmuller, and J. P. Johnson, Nucleic Acids Res. 18:1048, 1990). Nck was found to be constitutively phosphorylated on serine in resting NIH 3T3 cells. Platelet-derived growth factor (PDGF) treatment led to increased Nck phosphorylation on both tyrosine and serine. Nck was also found to be phosphorylated on tyrosine in epidermal growth factor (EGF)-treated A431 cells and in v-Src-transformed NIH 3T3 cells. Multiple sites of serine phosphorylation were detected in Nck from resting cells, and no novel sites were found upon PDGF or EGF treatment. A single major tyrosine phosphorylation site was found in Nck in both PDGF- and EGF-treated cells and in v-Src-transformed cells. This same tyrosine was phosphorylated in vitro by purified PDGF and EGF receptors and also by pp60c-src. We compared the phosphorylation of Nck and PLC-gamma 1 in several cell lines transformed by oncogenes with different modes of transformation. Although PLC-gamma 1 and Nck have significant amino acid identity, particularly in their SH3 regions, and both associate with growth factor receptors in a ligand-dependent manner, they were not always phosphorylated on tyrosine in a coincident manner.
Meisenhelder
J
J
Molecular Biology and Virology Laboratory, Salk Institute, San Diego, California 92186.
Hunter
T
T
D016475:3T3 Cells; D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002417:Cattle; D002455:Cell Division; D002460:Cell Line; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D015513:Oncogene Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D010449:Peptide Mapping; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D011233:Precipitin Tests; D051381:Rats; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism*; D010738:Type C Phospholipases / Q000276:immunology* / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D000911:Antibodies, Monoclonal; C078332:Nck protein; D015513:Oncogene Proteins; D010982:Platelet-Derived Growth Factor; D013438:Sulfhydryl Compounds; D014443:Tyrosine; D012694:Serine; D004815:Epidermal Growth Factor; D010738:Type C Phospholipases