LIGAND-INDUCED PHOSPHORYLATION OF A MURINE TUMOR SURFACE PROTEIN (TSP-180) ASSOCIATED WITH METASTATIC PHENOTYPE

  • 15 May 1989
    • journal article
    • research article
    • Vol. 49  (10) , 2615-2620
Abstract
A tumor surface protein (TSP-180) has been identified on murine lung carcinomas using two monoclonal antibodies (MoAbs) (135-13C and 346-11A). Quantitative analysis of TSP-180 on 3LL variants maintained either in vitro or in vivo indicates that TSP-180 is highly expressed in highly malignant metastatic cells. In reducing conditions, sodium dodecyl sulfate-polyacrylamide gel electrophoresis banding patterns of TSP-180 obtained with MoAb 135-13C from cell lysates of 3LL metastatic cells show three proteins migrating to Mr 204,000, 134,000, and 116,000. In the same experimental conditions MoAb 135-13C precipitates from low metastasizing ones only one band, corresponding to the lower molecular weight (Mr 116,000). All bands of TSP-180 observed in 3LL variants are labeled by lactoperoxidase-catalyzed radioiodination of viable cells, incorporate 32PO4, and contain carbohydrates, as judged by binding to wheat germ agglutinin. These results indicate that all proteins have external exposure on the cell surface and that at least some of TSP-180 proteins could be differentially regulated in different tumor cells (highly metastatic versus low metastatic). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis banding patterns and immunoblots obtained from cell lysates of 3LL variants by using a monoclonal antibody to phosphotyrosine (IG-2) indicate that this MoAb recognize proteins migrating with molecular weights identical to those reported for TSP-180. Moreover, the immunoblots of solubilized immunocomplex, obtained from cell lysates of 3LL variants by using MoAb 135-13C, demonstrate that MoAb IG-2 specifically reacts with TSP-180 proteins. Experiments undertaken in order to assess if some or all of TSP-180 protiens have tyrosine kinase activity demonstrate that MoAb 135-13C binding to the cell surface induces specific phosphorylation of the Mr 204,000 protein of TSP-180. Phosphoaminoacid analysis of the ligand-induced phosphorylated protein (pp204) demonstrates that this protein is phosphorylated at serine and tyrosine. Results reported lead us to hypothesize that TSP-180 is involved in growth-regulation mechanisms and that its high expression on cells with more malignant phenotype could be responsible for a proliferative advantage of such tumor clones.

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