Signaling at the inhibitory natural killer cell immune synapse regulates lipid raft polarization but not class I MHC clustering

Natural killer (NK) cell cytotoxicity is determined by a balance of positiv e and negative signals. Negative signals are transmitted by NK inhibitory r eceptors (killer immunoglobulin-like receptors, KIR) at the site of membran e apposition between an NK cell and a target cell, where inhibitory recepto rs become clustered with class I MHC ligands in an organized structure know n as an inhibitory NK immune synapse. Immune synapse formation in NK cells is poorly understood. Because signaling by NK inhibitory receptors could be involved in this process, the human NK tumor line YTS was transfected with signal-competent and signal-incompetent KIR2DL1. The latter were generated by truncating the KIR2DL1 cytoplasmic tail or by introducing mutations in the immunoreceptor tyrosine-based inhibition motifs. The KIR2DL1 mutants re tained their ability to cluster class I MHC ligands on NK cell interaction with appropriate target cells. Therefore, receptor-ligand clustering at the inhibitory NK immune synapse occurs independently of KIR2DL1 signal transd uction. However, parallel examination of NK cell membrane lipid rafts revea led that KIR2DL1 signaling is critical for blocking lipid raft polarization and NK cell cytotoxicity. Moreover, raft polarization was inhibited by rea gents that disrupt microtubules and actin filaments, whereas synapse format ion was not. Thus, NK lipid raft polarization and inhibitory NK immune syna pse formation occur by fundamentally distinct mechanisms.