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Abstract
Natural Killer (NK) cells act as early defenders against influenza infection by directly recognizing and responding to the presence of viral hemagglutinin, but are also capable of modulating the subsequent anti-viral CD8 T cell response. In these studies, we utilized an NK cell depletion model to investigate the contribution of NK cells to the generation and development of anti-influenza specific memory CD8 T cells. We found that the absence of NK cells during primary influenza infection resulted in increased numbers of influenza-specific memory CD8 T cells present in the lung and lung draining lymph node. Furthermore, this enlarged anti-influenza memory CD8 T cell pool was capable of mediating heterosubtypic protection against lethal viral challenge with fewer reactivated CD8 T cells infiltrating the respiratory tract, likely through increased early production of the anti-viral cytokines IFN- and TNF. Additionally, we utilized in vitro influenza infection and protein expression models to examine how hemagglutinin intrinsic features, such as receptor binding specificity and glycosylation levels, impact NK cell activation. We confirmed that influenza hemagglutinin can functionally activate NK cells in an NKp46 dependent manner. We demonstrated that human NKp46 preferentially binds to avian origin hemagglutinins when compared to those of human origin suggesting hemagglutinin sialic acid binding preference may contribute to NK cell activation. However, we also found evidence of previously undescribed sialic acid independent interactions mediating binding of NKp46 to influenza hemagglutinin. Lastly, we found that increasing the glycosylation level of influenza hemagglutinin led to decreased NKp46 binding suggesting that the natural accumulation of glycosylation sites by circulating influenza strains may act as a viral mechanism to evade NK cell activation. Together these data suggest that modulation of NK cell activation represents a novel method to improve anti-influenza memory CD8 T cell generation during vaccination and that mechanisms of NK cell activation should be considered in rational vaccine design.