Studies on the role of toll-like receptors in natural killer cell function

Abstract

Natural Killer (NK) cells express receptors that allow them to recognise pathogen and activate effector functions such as cytotoxicity and cytokine production. Among these receptors are the recently identified Toll-like receptors that recognise conserved pathogen structures and initiate innate immune responses. We demonstrate that human NK cells express TLR3, TLR7 and TLR8, and that these receptors are functional. TLR3 is expressed at the cell surface where it functions as a receptor for poly (I:C) in a lysosomal independent manner. TLR7/8 signalling is sensitive to chloroquine inhibition indicating a requirement for lysosomal signalling as for other cell types. Both R848, an agonist of human TLR7 and TLR8, and poly (I:C) activate NK cell cytotoxicity against Daudi target cells, which is partially IL-12 dependent. However, IFN-y production, another important effector function of NK cells in response to infection, is differentially regulated by these TLR agonists. In contrast to poly (I:C), R848 stimulates significant IFN-y production by NK cells. This is accessory cell dependent and was inhibited by addition of a neutralising anti-IL-12 antibody. Moreover, stimulation of purified monocyte populations with R848 results in IL-12 production and reconstitution of purified NK cells with monocytes results in increased IFN-gamma production in response to R848. We demonstrate that while NK cells constitutively express IRF-7, stimulation with R848 does not induce activation of IRF-7. Indeed, R848-induced NK cells cytotoxicity is IFN-a independent. However, manipulation of R848 for endosomal retention facilitates activation of IRF-7 in NK cells. In addition, we demonstrate that while resting NK cells do not respond directly to R848, they can be primed to do so by prior exposure to either IL-2 or IFN-a. Therefore, although NK cells can be directly activated by TLRs, accessory cells make a major contribution to effector functions such as IFN-y production and cytotoxicity. Direct sensing of viral products by NK cells through TLRs is a novel mechanism of pathogen recognition, and highlights the important role NK cells play in the immune response to viral infection.