Recombinant glycosyl-phosphatidylinositol-anchored proteins are not associated with protein kinases in transfected thymoma cells.
The cross-linking by antibody of some glycosyl-phosphatidyl-inositol (GPI)-anchored proteins on the plasma membrane of T cells leads to cell activation. Phosphorylation of proteins on tyrosine residues has a central role in the control of T cell activation, and non-receptor protein tyrosine kinases can be coprecipitated with immune complexes of GPI-anchored proteins in T cell lysates. In order to investigate the nature of this interaction, two recombinant GPI-anchored proteins were constructed (using the GPI signal sequence from Thy-1), and their associations with protein tyrosine kinases in stable transfectants of a mouse thymoma have been investigated. One recombinant GPI protein is the extracellular domain of the human complement receptor-1, normally an integral membrane protein, and the other is the secreted protein, human tissue inhibitor of metalloproteinases. The latter protein should be foreign to the cell surface and yet has been expressed as a GPI-anchored protein at levels equivalent to the highly expressed antigens Thy-1 and Ly6.A2 on mouse thymoma cells. Neither of the two recombinant proteins, when immunoprecipitated from NP40 lysates of transfected cells, was associated with protein tyrosine kinases in contrast with the natural endogenous GPI-anchored proteins Thy-1 and Ly6.A2 in non-transfected parental cells. Moreover, high expression of foreign recombinant GPI protein appears to interfere with the association of the natural GPI proteins with protein tyrosine kinases.
Clissold
P M
PM
Molecular Immunopathology Unit, MRC Centre, Cambridge, U.K.
D000818:Animals; D000951:Antigens, Neoplasm / Q000235:genetics / Q000378:metabolism; D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism; D005434:Flow Cytometry; D005960:Glucosides / Q000494:pharmacology; D006023:Glycoproteins / Q000235:genetics; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D051381:Rats; D017463:Receptors, Complement 3b / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D018800:Thy-1 Antigens / Q000235:genetics / Q000378:metabolism; D013945:Thymoma / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D013953:Thymus Neoplasms / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D019714:Tissue Inhibitor of Metalloproteinases; D014162:Transfection; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D005960:Glucosides; D006023:Glycoproteins; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D017463:Receptors, Complement 3b; D011994:Recombinant Proteins; D018800:Thy-1 Antigens; D019714:Tissue Inhibitor of Metalloproteinases; C018619:octyl-beta-D-glucoside; D011505:Protein-Tyrosine Kinases