Natural variation at position 45 in the D1 domain of lineage III killer cell immunoglobulin-like receptors (KIR) has major effects on the avidity and specificity for MHC class I

Abstract

Alternative lysine and methionine residues at position 44 in the D1 domain determine the specificities of human lineage III killer cell immunoglobulin-like receptors (KIR) for the C1 and C2 epitopes of HLA-C. KIR having glutamate 44 are also present in orangutans (Popy2DLB) and chimpanzees (Pt-2DL9) but notably absent from humans. Popy2DLB exhibits broad specificity for both the C1 and C2 epitopes, whereas Pt-2DL9 has narrow specificity for C2. Mutation of phenylalanine 45 in Popy2DLB to the cysteine residue present in Pt-2DL9 was sufficient to narrow the Popy2DLB specificity to be like that of Pt-2DL9. In contrast, replacement of cysteine 45 in Pt-2DL9 by phenylalanine had no effect on its C2 specificity, but reduced the avidity. In a similar manner, replacement of phenylalanine 45 with cysteine in Popy2DLA, which has lysine 44 and recognizes C1, maintained this specificity while reducing avidity. Position 45 is exceptionally variable, exhibiting twelve residues that distinguish KIR of different lineages and species. Our study demonstrates the potential for variation at position 45 to modulate KIR avidity and specificity for HLA-C. The various effects of position 45 mutation are consistent with a model in which a Popy2DLB-like receptor, having glutamate 44 and broad specificity for C1 and C2, facilitated the evolution of the C2 epitope from the C1 epitope and C2-specific KIR from C1-specific KIR. With the acquisition of C2 and C2-specific receptors, the selection against this broadly specific receptor led to its loss from the human line and narrowing of its specificity on the chimpanzee line.

Fig. 1

Alignment of positions of difference within the D1 and D2 domains of 13 human, chimpanzee and orangutan lineage III KIR. Popy2DLA is used as the consensus sequence, and identity with it is marked by a dot. Position 44 is marked in dark grey, and the residues selected for mutational analysis, positions 45, 70, 71, and 72, are marked in light grey

Fig. 2

Mutation at position 45 changes the specificity of Popy2DLB, while affecting only the avidity of Popy2DLA and Pt-2DL9. Shown are results of assays to measure the binding of nine KIR-Fc fusion proteins to beads coated with one of 18 HLA class I allotypes: eleven C1 and seven C2. Binding is shown for individual allotypes and for mean C1 and C2. Experiments were performed as described previously (Moesta et al. 2009; Older Aguilar et al. 2010). A minimum of three independent assays were performed for each fusion protein. Shown are the averaged data for all experiments