Regulation of immune responses by inhibitory receptors

National Institute Of Allergy And Infectious Diseases

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Abstract

Killer cell immunoglobulin-like receptors (KIR) that bind HLA-C block activation of human natural killer (NK) cells. This inhibition serves to protect healthy cells, which express HLA-C. At the same time, inhibition by KIR maintains NK cells in a state of high responsiveness to subsequent activation signals. This feature has been referred to as licensing of NK cells. Licensed NK cells are primed to kill cells that do not express HLA-C, i.e. that are missing self. We have shown that the inhibitory function of KIR is dependent on zinc, as Zn(II). Zinc induces the assembly of soluble KIR into filamentous polymers, as detected by electron microscopy. KIR filaments formed spontaneously, without addition of zinc, at functional inhibitory immunological synapses of NK cells with HLA-C+ cells. Live imaging was performed to visualize the distribution and movement of the zinc transporter ZnT1. ZnT1 is the main zinc channel at the plasma membrane for extrusion of zinc out of cells. A ZnT1 tagged with a fluorescent protein and expressed in an NK cell line remained evenly distributed at the plasma membrane of NK cells that formed activating immunological synapses with target cells. In contrast, ZnT1 moved towards NK-target cell contacts at inhibitory immunological synapses with HLA-C+ target cells, suggesting that it delivers the zinc required for inhibitory KIR function. Studies have shown that the low affinity interaction of KIR with HLA-C, and their dissociation rate, cannot explain the retention of KIR at immunological synapses, and that retention requires either tethering or confinement of KIR. Our hypothesis for the role of zinc in the inhibitory function of KIR is that zinc-dependent polymers reduce the lateral diffusion rate of KIR at the plasma membrane, thereby preventing rapid diffusion of KIR out of the synapse. We measured the diffusion of KIR tagged with a photo-switchable fluorescent protein, in the presence or absence of zinc in the culture medium. KIR diffusion was reduced after addition of zinc. The diffusion of a KIR mutant that fails to form zinc-dependent polymers was less sensitive to zinc. Similar measurements are being performed at inhibitory immunological synapses.