MHC class I-specific NK cell receptors inhibit natural killing, and presumably granule exocytosis, when engaged by target cell ligands but it is not yet known which specific activation receptor pathway for natural killing is inhibited or if these receptors influence other NK cell activities such as cytokine production. Moreover, an individual NK cell may express multiple inhibitory MHC class I receptors; these NK cell receptors may cooperate in inhibiting NK cell activity. To address these issues, we examined whether the murine Ly-49A NK cell receptor, specific for H-2D(d), can regulate granule exocytosis and NK cell cytokine responses. Expression of transfected H-2D(d) on tumor targets specifically inhibited granule release from Ly-49A+ NK cells. Importantly, Ly-49A engagement also inhibited target cell-induced cytokine (GM-CSF) secretion. Using a target cell-free system, we next determined that anti-Ly-49A mAb can regulate NK cell responses induced by specific stimuli. Cross-linking of NK1.1 with immobilized mAb induced granule release and TNF-α, IFN-γ, and GM-CSF secretion; all were inhibited by coimmobilized anti-Ly-49A. These effects were specific because an isotype- matched control mAb did not alter NK1.1-mediated responses. Therefore, these results demonstrate that the Ly-49A receptor can regulate granule exocytosis and cytokine secretion in response to targets and NK1.1 signaling, consistent with its function as an inhibitory receptor for MHC class I molecules in NK cell-mediated cytotoxicity. In addition, these results strongly support the thesis that signals transduced from Ly-49A alone are sufficient for mediating the inhibitory effects against target cells.
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