Abstract
Viral infections can affect the glycosylation pattern of glycoproteins involved in antiviral immunity. Given the importance of protein glycosylation for immune function, we investigated the effect that modulation of the highly conserved HLA class I N-glycan has on KIR:HLA interactions and NK cell function. We focused on HLA-B*57:01 and its interaction with KIR3DL1, which has been shown to play a critical role in determining the progression of a number of human diseases, including human immunodeficiency virus-1 infection. 721.221 cells stably expressing HLA-B*57:01 were treated with a panel of glycosylation enzyme inhibitors, and HLA class I expression and KIR3DL1 binding was quantified. In addition, the functional outcomes of HLA-B*57:01 N-glycan disruption/modulation on KIR3DL1ζ+ Jurkat reporter cells and primary human KIR3DL1+ NK cells was assessed. Different glycosylation enzyme inhibitors had varying effects on HLA-B*57:01 expression and KIR3DL1-Fc binding. The most remarkable effect was that of tunicamycin, an inhibitor of the first step of N-glycosylation, which resulted in significantly reduced KIR3DL1-Fc binding despite sustained expression of HLA-B*57:01 on 721.221 cells. This effect was paralleled by decreased activation of KIR3DL1ζ+ Jurkat reporter cells, as well as increased degranulation of primary human KIR3DL1+ NK cell clones when encountering HLA-B*57:01-expressing 721.221 cells that were pre-treated with tunicamycin. Overall, these results demonstrate that N-glycosylation of HLA class I is important for KIR:HLA binding and has an impact on NK cell function.
Highlights
Natural killer (NK) cells are part of the innate immune system, and serve as a first line of defense against intracellular pathogens and malignantly transformed cells
In order to examine the effect of different commercially available glycosylation inhibitors on killer cell immunoglobulin-like receptors (KIRs):human leukocyte antigen (HLA) interactions, we examined their effect on HLA-BÃ57:01 expression on HLA-BÃ57:01transduced 221 cells and on KIR3DL1-Fc binding to those cells
After preliminary testing of the effects that several different glycosylation inhibitors had on HLA class I expression and KIR binding, we decided to focus on TUN, the only inhibitor tested that did not decrease HLA class I surface expression but dramatically reduced KIR3DL1-Fc binding, and that had the unique function of completely inhibiting that addition of N-glycans
Summary
Natural killer (NK) cells are part of the innate immune system, and serve as a first line of defense against intracellular pathogens and malignantly transformed cells. HLA Glycosylation and KIR Binding intracellular bacteria [1,2,3,4]. NK cell activity is regulated via a number of activating and inhibitory receptors, including killer cell immunoglobulin-like receptors (KIRs) [1, 5], which recognize human leukocyte antigen (HLA) proteins. While binding of inhibitory KIRs to HLA class I molecules results in NK cell inhibition, engagement of activating KIRs or loss of inhibitory KIR binding can lead to NK cell activation [8, 9]. The KIR-HLA interaction has been demonstrated to have an important impact on viral infections, including human immunodeficiency virus-1 (HIV-1). In HIV-1 positive individuals, KIR3DL1+ NK cells expanded preferentially in the presence of HLA-Bw4-80I, the ligand class for KIR3DL1 [10, 11]
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