Genistein, a tyrosine kinase inhibitor, decreased the affinity of p56lck to beta-chain of interleukin-2 receptor in human natural killer (NK)-rich cells and decreased NK-mediated cytotoxicity.
Intracellular signal transduction has been reported to be triggered by phosphorylation of interleukin-2 (IL-2) receptor by IL-2. In order to clarify the effect of tyrosine phosphorylation of IL-2 receptors on cell-mediated cytotoxicity by natural killer (NK) cells, we studied the effect of a tyrosine kinase inhibitor, genistein, on the lethal effects of NK-rich cells for K562 cells. Exposure of NK-rich cells to IL-2 (100 U/ml) for 3 days increased their cytotoxicities against K562 cells. The effect was reduced in the presence of 10 micrograms/ml of genistein. Samples immunoprecipitated by anti-IL-2R beta antibodies were prepared from NK-rich fractions with or without exposure to IL-2 and/or genistein. Coprecipitated proteins with 75, 65, and 56 kDa were detected with an antiphosphotyrosine antibody. The amount of phosphorylated tyrosine residues of 56-kDa protein, which was predominantly detected in NK-rich cells, was remarkably increased by IL-2 treatment. The enhanced phosphorylation of 56-kDa protein was reduced by the presence of genistein. These results suggested that IL-2 increased tyrosine phosphorylation and the affinity to IL-2R beta of the 56-kDa proteins in NK-rich cells. Immunoprecipitated samples by anti-IL-2R beta were reblotted with anti-p56lck antibody and revealed that the 56-kDa protein was identified to be p56lck. The increase of coprecipitated p56lck with anti-IL-2R beta antibody by the treatment with IL-2 suggested that the affinity of p56lck to IL-2R beta was increased by IL-2 in NK-rich cells. The amount of coprecipitated p56lck with IL-2R beta was reduced in the sample exposed to genistein. The affinity of p56lck to IL-2R beta was considered to be regulated by IL-2-induced tyrosine phosphorylation. Our results demonstrated a potential role for tyrosine kinase, p56lck, in the signaling events that regulate the cytotoxicity by NK-rich cells.
Nishio
K
K
Pharmacology Division, National Cancer Center Research Institute, Tokyo, Japan.
Miura
K
K
Ohira
T
T
Heike
Y
Y
Saijo
N
N
D003602:Cytotoxicity, Immunologic / Q000187:drug effects*; D019833:Genistein; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology; D007529:Isoflavones / Q000494:pharmacology*; D007694:Killer Cells, Natural / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D015375:Receptors, Interleukin-2 / Q000378:metabolism*
D007376:Interleukin-2; D007529:Isoflavones; D011518:Proto-Oncogene Proteins; D015375:Receptors, Interleukin-2; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck)