Peptide-Dependent Recognition of HLA-B*57:01 by KIR3DS1.
ABSTRACT: UNLABELLED:Killer cell immunoglobulin-like receptors (KIRs) play an important role in the activation of natural killer (NK) cells, which in turn contribute to the effective immune control of many viral infections. In the context of HIV infection, the closely related KIR3DL1 and KIR3DS1 molecules, in particular, have been associated with disease outcome. Inhibitory signals via KIR3DL1 are disrupted by downregulation of HLA class I ligands on the infected cell surface and can also be impacted by changes in the presented peptide repertoire. In contrast, the activatory ligands for KIR3DS1 remain obscure. We used a structure-driven approach to define the characteristics of HLA class I-restricted peptides that interact with KIR3DL1 and KIR3DS1. In the case of HLA-B*57:01, we used this knowledge to identify bona fide HIV-derived peptide epitopes with similar properties. Two such peptides facilitated productive interactions between HLA-B*57:01 and KIR3DS1. These data reveal the presence of KIR3DS1 ligands within the HIV-specific peptide repertoire presented by a protective HLA class I allotype, thereby enhancing our mechanistic understanding of the processes that enable NK cells to impact disease outcome. IMPORTANCE:Natural killer (NK) cells are implicated as determinants of immune control in many viral infections, but the precise molecular mechanisms that initiate and control these responses are unclear. The activating receptor KIR3DS1 in combination with HLA-Bw4 has been associated with better outcomes in HIV infection. However, evidence of a direct interaction between these molecules is lacking. In this study, we demonstrate that KIR3DS1 recognition of HLA-Bw4 is peptide dependent. We also identify HIV-derived peptide epitopes presented by the protective HLA-B*57:01 allotype that facilitate productive interactions with KIR3DS1. Collectively, these findings suggest a mechanism whereby changes in the peptide repertoire associated with viral infection provide a trigger for KIR3DS1 engagement and NK cell activation.
Project description:NK cell activity is regulated by the integration of positive and negative signals. One important source of these signals for human NK cells is the killer Ig-like receptor (KIR) family, which includes both members that transduce positive and those that generate negative signals. KIR3DL1 inhibits NK cell activity upon engagement by its ligand HLA-Bw4. The highly homologous KIR3DS1 is an activating receptor, which is implicated in the outcome of a variety of pathological situations. However, unlike KIR3DL1, direct binding of KIR3DS1(+) cells to HLA has not been demonstrated. We analyzed four key amino acid differences between KIR3DL1*01502 and KIR3DS1*013 to determine their role in KIR binding to HLA. Single substitutions of these residues dramatically reduced binding by KIR3DL1. In the reciprocal experiment, we found that the rare KIR3DS1 allotype KIR3DS1*014 binds HLA-Bw4 even though it differs from KIR3DS1*013 at only one of these positions (position 138). This reactivity was unexpectedly dependent on residues at other variable positions, as HLA-Bw4 binding was lost in receptors with KIR3DL1-like residues at both positions 199 and 138. These data provide the first evidence, to our knowledge, for the direct binding of KIR3DS1(+) cells to HLA-Bw4 and highlight the key role for position 138 in determining ligand specificity of KIR3DS1. They also reveal that KIR3DS1 reactivity and specificity is dictated by complex interactions between the residues in this region, suggesting a unique functional evolution of KIR3DS1 within the activating KIR family.
Project description:Natural killer (NK) cells play a role in the clearance of viral infections. Combinations of alleles at the polymorphic HLA-B locus and the NK cell surface killer immunoglobulin-like receptor locus KIR3DL1/S1 have been shown to influence time to AIDS in HIV-infected individuals and risk of seroconversion in HIV exposed seronegative (HESN) subjects. Here, we assessed time to seroconversion or duration of seronegative status in a group of 168 HIV exposed individuals, including 74 seroconverters and 94 HESN based on carriage or not of KIR3DL1/S1/HLA-B genotypes previously shown to be associated with protection from infection and/or slow time to AIDS. KIR3DL1/S1 genotyping was performed by sequence-specific primer polymerase chain reaction using two pairs of specific primers for each locus. The MHC class IB locus was typed to four-position resolution to resolve Bw4 and Bw6 alleles and the amino acid present at position 80. KIR3DL1/S1 heterozygotes became HIV infected significantly faster than KIR3DS1 homozygotes. Individuals who carried both KIR3DS1 and Bw4*80I did not remain HIV seronegative longer than those from a control group who were homozygous for HLA-Bw6 and carried no HLA-A locus Bw4 alleles Subjects who were *h/*y+B*57 showed a trend towards slower time to serconversion than those with other KIR3DL1 homozygous and KIR3DL1/S1 heterozygous genotypes. Thus, KIR3DS1 homozygosity is associated with protection from HIV infection while co-carriage of KIR3DS1 and Bw4*80I is not. The requirements for protection from HIV infection can differ from those that influence time to AIDS in HIV infected individuals.
Project description:Killer Immunoglobulin-like Receptors (KIR) are a family of receptors expressed on natural killer (NK) and T-cell subsets. KIR3DL1 is a highly polymorphic receptor that binds to groups of HLAA and HLA-B allotypes that express the Bw4 epitope. The variation in KIR3DL1 allotypes manifests at a number of levels. Most dramatically, a common allelic variant encodes an activating rather than an inhibitory receptor (KIR3DS1). In addition, sequence variants can affect both the frequency of expression within the NK cell population and the intensity of expression on a given cell. KIR3DL1 polymorphism also influences the interaction with HLA-Bw4 molecules, due to contacts with the HLA molecule itself and sensitivity to the presented peptide. A body of evidence from genetic association studies supports the biological significance not only of the interaction of KIR3DL1 with HLA-Bw4 but also the functional variation seen with different KIR3DL1 and HLA allotypes. In this review, we discuss our current understanding of KIR3DL1 function and our recent insights from the structure of the KIR3DL1 in complex with HLA. In addition, we will summarize our current understanding of KIR3DS1, including its ligand specificity and its role in immune responses.
Project description:A genome-wide screen for large structural variants showed that a copy number variant (CNV) in the region encoding killer cell immunoglobulin-like receptors (KIR) associates with HIV-1 control as measured by plasma viral load at set point in individuals of European ancestry. This CNV encompasses the KIR3DL1-KIR3DS1 locus, encoding receptors that interact with specific HLA-Bw4 molecules to regulate the activation of lymphocyte subsets including natural killer (NK) cells. We quantified the number of copies of KIR3DS1 and KIR3DL1 in a large HIV-1 positive cohort, and showed that an increase in KIR3DS1 count associates with a lower viral set point if its putative ligand is present (p = 0.00028), as does an increase in KIR3DL1 count in the presence of KIR3DS1 and appropriate ligands for both receptors (p = 0.0015). We further provide functional data that demonstrate that NK cells from individuals with multiple copies of KIR3DL1, in the presence of KIR3DS1 and the appropriate ligands, inhibit HIV-1 replication more robustly, and associated with a significant expansion in the frequency of KIR3DS1+, but not KIR3DL1+, NK cells in their peripheral blood. Our results suggest that the relative amounts of these activating and inhibitory KIR play a role in regulating the peripheral expansion of highly antiviral KIR3DS1+ NK cells, which may determine differences in HIV-1 control following infection.
Project description:Several studies described an association between killer-cell immunoglobulin-like receptor (KIR)/HLA gene combinations and clinical outcomes in various diseases. In particular, an important combined role for KIR3DS1 and HLA-B Bw4-I80 in controlling viral infections and a higher protection against leukemic relapses in donor equipped with activating KIRs in haplo-HSCT has been described. Here, we show that KIR3DS1 mediates positive signals upon recognition of HLA-B*51 (Bw4-I80) surface molecules on target cells and that this activation occurs only in Bw4-I80neg individuals, including those carrying particular KIR/HLA combination settings. In addition, killing of HLA-B*51 transfected target cells mediated by KIR3DS1+/NKG2A+ natural killer (NK) cell clones from Bw4-I80neg donors could be partially inhibited by antibody-mediated masking of KIR3DS1. Interestingly, KIR3DS1-mediated recognition of HLA-B*51 could be better appreciated under experimental conditions in which the function of NKG2D was reduced by mAb-mediated blocking. This experimental approach may mimic the compromised function of NKG2D occurring in certain viral infections. We also show that, in KIR3DS1+/NKG2A+ NK cell clones derived from an HLA-B Bw4-T80 donor carrying 2 KIR3DS1 gene copy numbers, the positive signal generated by the engagement of KIR3DS1 by HLA-B*51 resulted in a more efficient killing of HLA-B*51-transfected target cells. Moreover, in these clones, a direct correlation between KIR3DS1 and NKG2D surface density was detected, while the expression of NKp46 was inversely correlated with that of KIR3DS1. Finally, we analyzed KIR3DS1+/NKG2A+ NK cell clones from a HLA-B Bw4neg donor carrying cytoplasmic KIR3DL1. Although these clones expressed lower levels of surface KIR3DS1, they displayed responses comparable to those of NK cell clones derived from HLA-B Bw4neg donors that expressed surface KIR3DL1. Altogether these data suggest that, in particular KIR/HLA combinations, KIR3DS1 may play a role in the process of human NK cell education.
Project description:The function of natural killer (NK) cells is controlled by several activating and inhibitory receptors, including the family of killer-immunoglobulin-like receptors (KIRs). One distinctive feature of KIRs is the extensive number of various haplotypes generated by the gene content within the KIR gene locus as well as by highly polymorphic members of the KIR gene family, namely KIR3DL1/S1. Within the KIR3DL1/S1 gene locus, KIR3DS1 represents a conserved allelic variant and displays other unique features in comparison to the highly polymorphic KIR3DL1 allele. KIR3DS1 is present in all human populations and belongs to the KIR haplotype group B. KIR3DS1 encodes for an activating receptor featuring the characteristic short cytoplasmic tail and a positively charged residue within the transmembrane domain, which allows recruitment of the ITAM-bearing adaptor molecule DAP12. Although HLA class I molecules are thought to represent natural KIR ligands, and HLA-Bw4 molecules serve as ligands for KIR3DL1, the ligand for KIR3DS1 still needs to be identified. Despite the lack of formal evidence for an interaction of KIR3DS1 with HLA-Bw4-I80 or any other HLA class I subtype to date, a growing number of associations between the presence of KIR3DS1 and the outcome of viral infections have been described. Especially, the potential protective role of KIR3DS1 in combination with HLA-Bw4-I80 in the context of HIV-1 infection has been studied intensively. In addition, a number of recent studies have associated the presence or absence of KIR3DS1 with the occurrence and outcome of some malignancies, autoimmune diseases, and graft-versus-host disease (GVHD). In this review, we summarize the present knowledge regarding the characteristics of KIRD3S1 and discuss its role in various human diseases.
Project description:Variable interaction between the Bw4 epitope of HLA-B and the polymorphic KIR3DL1/S1 system of inhibitory and activating NK cell receptors diversifies the development, repertoire formation, and response of human NK cells. KIR3DL1*004, a common KIR3DL1 allotype, in combination with Bw4(+) HLA-B, slows progression of HIV infection to AIDS. Analysis in this study of KIR3DL1*004 membrane traffic in NK cells shows this allotype is largely misfolded but stably retained in the endoplasmic reticulum, where it binds to the chaperone calreticulin and does not induce the unfolded protein response. A small fraction of KIR3DL1*004 folds correctly and leaves the endoplasmic reticulum to be expressed on the surface of primary NK and transfected NKL cells, in a form that can be triggered to inhibit NK cell activation and secretion of IFN-?. Consistent with this small proportion of correctly folded molecules, trace amounts of MHC class I coimmunoprecipitated with KIR3DL1*004. There was no indication of any extensive intracellular interaction between unfolded KIR3DL1*004 and cognate Bw4(+) HLA-B. A similarly limited interaction of Bw4 with KIR3DL1*002, when both were expressed by the same cell, was observed despite the efficient folding of KIR3DL1*002 and its abundance on the NK cell surface. Several positions of polymorphism modulate KIR3DL1 abundance at the cell surface, differences that do not necessarily correlate with the potency of allotype function. In this context, our results suggest the possibility that the effect of Bw4(+) HLA-B and KIR3DL1*004 in slowing progression to AIDS is mediated by interaction of Bw4(+) HLA-B with the small fraction of cell surface KIR3DL1*004.
Project description:BACKGROUND: Natural killer (NK) cells have emerged as pivotal players in innate immunity, especially in the defense against viral infections and tumors. Killer immunoglobulin-like receptors (KIRs)--an important recognition receptor expressed on the surface of NK cells--regulate the inhibition and/or activation of NK cells after interacting with human leukocyte antigen (HLA) class I ligands. Various KIR genes might impact the prognosis of many different diseases. The implications of KIR-HLA interaction in HIV disease progression remains poorly understood. METHODS: Here, we studied KIR genotypes, mRNA levels, HLA genotypes, CD4+ T cell counts and viral loads in our cohort of Human Immunodeficiency Virus (HIV)-infected individuals, a group that includes HIV long-term nonprogressors (LTNPs) and typical progressors (TPs). RESULTS: We found that the frequency of KIR3DS1/L1 heterozygotes with HLA-Bw4-80I gene was much higher in LTNPs than in TPs (P?=?0.001) and that the KIR3DL1 homozygotes without HLA-Bw4-80I gene had higher viral loads and lower CD4+ T cell counts (P?=?0.014 and P?=?0.021, respectively). Our study also confirmed that homozygosity for the HLA-Bw6 allele was associated with rapid disease progression. In addition to the aforementioned results on the DNA level, we observed that higher level expression of KIR3DS1 mRNA was in LTNP group, and that higher level expression of KIR3DL1 mRNA was in TP group. CONCLUSIONS: Our data suggest that different KIR-HLA genotypes and different levels of transcripts associate with HIV disease progression.
Project description:Killer Ig-like receptors (KIRs) control the activation of human NK cells via interactions with peptide-laden HLAs. KIR3DL1 is a highly polymorphic inhibitory receptor that recognizes a diverse array of HLA molecules expressing the Bw4 epitope, a group with multiple polymorphisms incorporating variants within the Bw4 motif. Genetic studies suggest that KIR3DL1 variation has functional significance in several disease states, including HIV infection. However, owing to differences across KIR3DL1 allotypes, HLA-Bw4, and associated peptides, the mechanistic link with biological outcome remains unclear. In this study, we elucidated the impact of KIR3DL1 polymorphism on peptide-laden HLA recognition. Mutational analysis revealed that KIR residues involved in water-mediated contacts with the HLA-presented peptide influence peptide binding specificity. In particular, residue 282 (glutamate) in the D2 domain underpins the lack of tolerance of negatively charged C-terminal peptide residues. Allotypic KIR3DL1 variants, defined by neighboring residue 283, displayed differential sensitivities to HLA-bound peptide, including the variable HLA-B*57:01-restricted HIV-1 Gag-derived epitope TW10. Residue 283, which has undergone positive selection during the evolution of human KIRs, also played a central role in Bw4 subtype recognition by KIR3DL1. Collectively, our findings uncover a common molecular regulator that controls HLA and peptide discrimination without participating directly in peptide-laden HLA interactions. Furthermore, they provide insight into the mechanics of interaction and generate simple, easily assessed criteria for the definition of KIR3DL1 functional groupings that will be relevant in many clinical applications, including bone marrow transplantation.
Project description:KIR3DL1 is a polymorphic inhibitory receptor that modulates NK cell activity through interacting with HLA-A and HLA-B alleles that carry the Bw4 epitope. Amino acid polymorphisms throughout KIR3DL1 impact receptor surface expression and affinity for HLA. KIR3DL1/S1 encodes inhibitory and activating alleles, but despite high homology with KIR3DL1, the activating receptor KIR3DS1 does not bind the same ligand. Allele KIR3DL1*009 resulted from a gene recombination event between the inhibitory receptor allele KIR3DL1*001 and the activating receptor allele KIR3DS1*013. This study analyzed the functional impact of KIR3DS1-specific polymorphisms on KIR3DL1*009 surface expression, binding to HLA, and functional capacity. Flow-cytometric analysis of primary human NK cells as well as transfected HEK293T cells shows that KIR3DL1*009 is expressed at a significantly lower surface density compared with KIR3DL1*001. Using recombinant proteins of KIR3DL1*001, KIR3DL1*009, and KIR3DS1*013 to analyze binding to HLA, we found that although KIR3DL1*009 displayed some evidence of binding to HLA compared with KIR3DS1*013, the binding was minimal compared with KIR3DL1*001 and KIR3DL1*005. Mutagenesis of polymorphic sites revealed that the surface phenotype and reduced binding of KIR3DL1*009 are caused by the combined amino acid polymorphisms at positions 58 and 92 within the D0 extracellular domain. Resulting from these effects, KIR3DL1*009(+) NK cells exhibited significantly less inhibition by HLA-Bw4(+) target cells compared with KIR3DL1*001(+) NK cells. The data from this study contribute novel insight into how KIR3DS1-specific polymorphisms in the extracellular region impact KIR3DL1 surface expression, ligand binding, and inhibitory function.