Spontaneous clustering and tyrosine phosphorylation of NK cell inhibitory receptor induced by ligand binding.
Inhibition of NK cell cytotoxicity by killer cell Ig-like receptors (KIR) depends on phosphorylation of cytoplasmic tyrosines in KIR, which recruit tyrosine phosphatase Src homology protein tyrosine phosphatase 1. It is not clear how KIR, whose function lies downstream of a tyrosine kinase, succeeds in blocking proximal NK cell activation signals upon binding HLA class I on target cells. Here we show that mixing NK cells with insect cells expressing HLA-C was sufficient to induce clustering of KIR, and phosphorylation of KIR and SHP-1. Transient phosphorylation of KIR was detected in the presence of pervanadate, an inhibitor of protein tyrosine phosphatases, at suboptimal concentration. Phosphorylation of KIR was specifically induced by ligand binding because it was detected only when HLA-C was loaded with a peptide that permits KIR binding. KIR phosphorylation was not dependent on ICAM-1-mediated adhesion and was not blocked by inhibition of actin polymerization, but required Zn(2+). Fluorescence resonance energy transfer between HLA-C molecules revealed close molecular interactions induced by KIR binding. These results demonstrate tight clustering of KIR and rapid KIR phosphorylation induced simply by binding to HLA-C. The unique property of KIR to become phosphorylated in the absence of adhesion and of actin cytoskeleton rearrangement explains how KIR can efficiently block early activation signals during NK-target cell contacts.
Faure
Mathias
M
Laboratory of Immunogenetics, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20852, USA.
Barber
Domingo F
DF
Takahashi
Stephenie M
SM
Jin
Tian
T
Long
Eric O
EO
D000818:Animals; D002448:Cell Adhesion / Q000276:immunology; D002460:Cell Line; D002461:Cell Line, Transformed; D002999:Clone Cells; D004330:Drosophila; D031541:Fluorescence Resonance Energy Transfer; D015236:HLA-C Antigens / Q000378:metabolism / Q000502:physiology; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000502:physiology; D007694:Killer Cells, Natural / Q000378:metabolism*; D008024:Ligands; D010766:Phosphorylation; D011485:Protein Binding / Q000276:immunology; D011940:Receptor Aggregation / Q000276:immunology*; D011971:Receptors, Immunologic / Q000378:metabolism*; D054340:Receptors, KIR; D054342:Receptors, KIR2DL1; D014162:Transfection; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000378:metabolism*; D015032:Zinc / Q000502:physiology; D019061:src-Family Kinases / Q000502:physiology
D015236:HLA-C Antigens; C038653:HLA-C*03 antigen; C039966:HLA-C*04 antigen; D008024:Ligands; D011971:Receptors, Immunologic; D054340:Receptors, KIR; D054342:Receptors, KIR2DL1; D018799:Intercellular Adhesion Molecule-1; D014443:Tyrosine; D019061:src-Family Kinases; D015032:Zinc