Abstract
Natural killer (NK) cells are key effectors of innate immunity and contribute to early control of viral infections. Their activity is regulated in part by interactions between killer cell immunoglobulin-like receptors (KIRs) on NK cells and major histocompatibility complex class I (MHC I) molecules on target cells. To investigate how MHC I-bound peptides influence these interactions in the context of simian immunodeficiency virus (SIV) infection of rhesus macaques, we employed ultradense peptide arrays and screened pooled matrices of overlapping peptides to identify sequences presented by common MHC I ligands of rhesus macaque KIRs. We identified over 20 distinct SIV peptides bound by Mamu-A3*13 and -B*041, both MHC I molecules that interact with high-frequency KIR allotypes. To assess peptide-specific effects on KIR-MHC I interactions, TAP-deficient, MHC I-transduced 721.221 cells were incubated with SIV-derived peptides and stained with KIR-IgG-Fc fusion proteins. KIR engagement was corroborated using a reporter cell line expressing chimeric KIR-CD3ζ receptors. Functional impacts of bound viral peptide sequences were confirmed in cytotoxicity assays using primary NK cells. Several peptides enhanced or diminished KIR engagement despite stabilizing similar MHC I surface levels, indicating peptide-dependent modulation of KIR binding. Notably, single amino acid substitutions at the penultimate position of peptides bound by Mamu-B*041 and Mamu-A3*13 significantly altered binding by KIR3DL01 and KIR3DL05, underscoring the importance of this residue for KIR engagement. These findings advance our understanding of KIR-MHC I interactions in the rhesus macaque and illustrate how variation in MHC I-bound peptides may influence NK cell responses.