Gs is the major G protein involved in interleukin-2-activated natural killer (IANK) cell-mediated cytotoxicity. Successful introduction of anti-G protein antibodies inside streptolysin O-permeabilized IANK cells.
In the present work, anti-G protein antibodies were introduced inside streptolysin permeabilized O interleukin-2-activated natural killer (IANK) cells. Successful entry of the antibodies was determined by flow cytometry and fluorescence microscopy. Permeabilized cells showed typical large granular lymphocyte morphology and remained functional, significantly lysing both NK-sensitive K562 cells and NK-resistant/IANK-sensitive RAJI target cells. This method was utilized to study the effect of anti-G protein antibodies on the functional activities of IANK cells. Anti-Gs antibody inhibited IANK cell killing of RAJI but not of K562 target cells. Further analysis showed that K562 and RAJI cells enhance the binding of guanosine 5'-O-(thiotriphosphate) to IANK cell membranes, and increase the hydrolysis of [32P]GTP in these membranes. Immunoblot analysis showed that K562 and RAJI cells induce the release of alpha o, but not alpha i, alpha s, or alpha q.11 from IANK cell membranes. Cumulatively, these data indicate that putative receptors recognizing K562 or RAJI target cells are coupled to Go in IANK cells, however, only Gs seems to be coupled to receptors recognizing RAJI target cells. Our results point out the importance of Gs protein as a mediator of cellular cytotoxicity of the anti-tumor effector cells.
Maghazachi
A A
AA
Northeastern Ontario Regional Cancer Centre, Laurentian University, Sudbury, Ontario, Canada.
Al-Aoukaty
A
A
D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D001426:Bacterial Proteins; D002051:Burkitt Lymphoma; D002460:Cell Line; D002463:Cell Membrane Permeability; D003602:Cytotoxicity, Immunologic; D005434:Flow Cytometry; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000276:immunology / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000494:pharmacology; D007376:Interleukin-2 / Q000276:immunology* / Q000494:pharmacology; D007694:Killer Cells, Natural / Q000187:drug effects / Q000276:immunology / Q000502:physiology*; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive; D008213:Lymphocyte Activation; D011817:Rabbits / Q000276:immunology; D013301:Streptolysins; D014407:Tumor Cells, Cultured
D000906:Antibodies; D001426:Bacterial Proteins; D007074:Immunoglobulin G; D007376:Interleukin-2; D013301:Streptolysins; C025964:streptolysin O; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins