(471) Constant engagement of a natural killer (NK) cell activation receptor with self-MHC-I renders hyporesponsive NK cells

Introduction/Rationale: Activating NK cell receptors are critical for controlling acute murine cytomegalovirus (MCMV) infection, most clearly demonstrated in C57BL/6 mice, where resistance is mediated by the Ly49H receptor through recognition of the viral ligand m157 expressed on infected cells. NK cell–dependent resistance to MCMV is also observed in Ly49H-deficient strains, such as MA/My, where Ly49P and Ly49R can recognize MCMV-infected cells in a class I MHC–dependent manner in vitro. However, the role of these receptors in vivo has not been explored. In addition, these activating receptors may engage self–MHC class I ligands in vivo, yet the physiological relevance of this interaction remains poorly defined.

Methods: To selectively assess Ly49P function in vivo, we generated Ly49- and MHC class I–deficient mice expressing a single Ly49P receptor on NK cells by knocking in Ly49P cDNA into the Ncr1 locus. Founder mice were then subsequently crossed onto distinct MHC class I backgrounds.

Results: NK cells from Ly49P-KkDk or Ly49P-KdDd mice exhibited reduced surface expression of Ly49P compared with Ly49P-KbDb mice, suggesting MHC class I–dependent masking of Ly49P. Consistently, Ly49P-KkDk NK cells showed diminished degranulation and IFN-γ production compared with controls following anti-Ly49P or NK1.1 stimulation, indicating a generalized hyporesponsive state to ITAM-associated signaling. This anergic state appeared to be regulated at the level of proximal signaling, as NK cell responsiveness was restored by PMA/ionomycin stimulation, which bypasses early signaling events. Notably, disruption of Ly49P–MHC class I cis interactions by citric acid treatment restored responsiveness of hyporesponsive NK cells to both Ly49P and NK1.1 crosslinking.

Conclusion: Thus, sustained stimulation of a self-specific activating receptor in vivo induces a hyporesponsive state to stimulation through both cognate and unrelated activating receptors.