Peptide selectivity discriminates NK cells from KIR2DL2‐ and KIR2DL3‐positive individuals

Natural killer cells are controlled by peptide selective inhibitory receptors for MHC class I, including the killer cell immunoglobulin‐like receptors (KIRs). Despite having similar ligands, KIR2DL2 and KIR2DL3 confer different levels of protection to infectious disease. To investigate how changes in peptide repertoire may differentially affect NK cell reactivity, NK cells from KIR2DL2 and KIR2DL3 homozygous donors were tested for activity against different combinations of strong inhibitory (VAPWNSFAL), weak inhibitory (VAPWNSRAL), and antagonist peptide (VAPWNSDAL). KIR2DL3‐positive NK cells were more sensitive to changes in the peptide content of MHC class I than KIR2DL2‐positive NK cells. These differences were observed for the weakly inhibitory peptide VAPWNSRAL in single peptide and double peptide experiments (p < 0.01 and p < 0.03, respectively). More significant differences were observed in experiments using all three peptides (p < 0.0001). Mathematical modeling of the experimental data demonstrated that VAPWNSRAL was dominant over VAPWNSFAL in distinguishing KIR2DL3‐ from KIR2DL2‐positive donors. Donors with different KIR genotypes have different responses to changes in the peptide bound by MHC class I. Differences in the response to the peptide content of MHC class I may be one mechanism underlying the protective effects of different KIR genes against infectious disease.     

Increased frequency and function of KIR2DL1–3+ NK cells in primary HIV‐1 infection are determined by HLA‐C group haplotypes

The acquisition and maintenance of NK‐cell function is mediated by inhibitory killer‐cell immunoglobulin‐like receptors (KIRs) through their interaction with HLA class I molecules. Recently, HLA‐C expression levels were shown to be correlated with protection against multiple outcomes of HIV‐1 infection; however, the underlying mechanisms are poorly understood. As HLA‐C is the natural ligand for the inhibitory receptors KIR2DL1 and KIR2DL2/3, we sought to determine whether HLA‐C group haplotypes affect NK‐cell responses during primary HIV‐1 infection. The phenotypes and functional capacity of NK cells derived from HIV‐1‐positive and HIV‐1‐negative individuals were assessed (N = 42 and N = 40, respectively). HIV‐1 infection was associated with an increased frequency of KIR2DL1–3⁺ NK cells. Further analysis showed that KIR2DL1⁺ NK cells were selectively increased in individuals homozygous for HLA‐C2, while HLA‐C1‐homozygous individuals displayed increased proportions of KIR2DL2/3⁺ NK cells. KIR2DL1–3⁺ NK cells were furthermore more polyfunctional during primary HIV‐1 infection in individuals also encoding for their cognate HLA‐C group haplotypes, as measured by degranulation and IFN‐γ and TNF‐α production. These results identify a novel relationship between HLA‐C and KIR2DL⁺ NK‐cell subsets and demonstrate that HLA‐C‐mediated licensing modulates NK‐cell responses to primary HIV‐1 infection.     

Homogenous expression of killer cell immunoglobulin-like receptors (KIR) on polyclonal natural killer cells detected by a monoclonal antibody to KIR2D

The activity of human natural killer (NK) cells is in part regulated by the expression of killer cell immunoglobulin (Ig)-like receptors (KIR) that recognize major histocompatibility complex (MHC) class I and can inhibit NK cell cytotoxicity. A monoclonal anti-KIR antibody was established and designated Lig1. Lig1 was shown to be specific for KIR in cell-surface staining and to react with all KIR2D, except KIR2DL4 which lacks a D1 domain, but not with KIR3D molecules in an enzyme-linked immunoadsorbent assay (ELISA) and Western blotting. Unlike other anti-KIR antibodies, Lig1 did not inhibit binding of KIR-Ig-fusion proteins to MHC-class I expressing cells nor did it interfere with KIR-mediated inhibition of NK cell cytotoxicity in a functional assay. Lig1 reacted with all NK cells in polyclonal NK populations from different donors, demonstrating that all NK cells express at least one KIR2D receptor.     

A vision of KIR variation at super resolution

The Ninth Killer Cell Immunoglobulin‐Like Receptor (KIR) Workshop was held in Winchester, UK late in the summer of 2015. The extraordinary diversity of KIR and its functional consequences were key themes throughout the meeting. Novel sequencing technologies and new bioinformatics techniques continue to increase our understanding of the genetic diversity and evolution of the KIR; while a deeper understanding of KIR functions, including their specificity for MHC and its peptide ligands, are generating more refined models of their role in disease. Limited to 100 delegates from around the world, this intimate workshop facilitated vigorous discussion, generating new ideas for research in this ever‐expanding field.     

Increased killer immunoglobulin-like receptor expression and functional defects in natural killer cells in lung cancer

Frequencies of natural killer (NK) cells from patients with non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC) did not differ from healthy controls. A higher proportion of NK cells from NSCLC patients expressed the killer immunoglobulin-like receptor (KIR) CD158b than in controls (P = 0.0004), in the presence or absence of its ligand, HLA-C1. A similar result was obtained for CD158e in the presence of its ligand HLA-Bw4 in NSCLC patients (P = 0.003); this was entirely attributable to the Bw4I group of alleles in the presence of which a fivefold higher percentage of CD158e(+) NK cells was found in NSCLC patients than controls. Proportions of CD158b(+) NK cells declined with advancing disease in NSCLC patients. Expression of NKp46, CD25 and perforin A, and production of interferon-γ following stimulation with interleukin-12 and interleukin-18, were all significantly lower in NK cells from NSCLC patients than in controls. Both NK cell cytotoxicity and granzyme B expression were also reduced in lung cancer patients. Increased inhibitory KIR expression would decrease NK cell cytotoxic function against tumour cells retaining class I HLA expression. Furthermore, the reduced ability to produce interferon-γ would restrict the ability of NK cells to stimulate T-cell responses in patients with lung cancer.     

Deciphering the killer‐cell immunoglobulin‐like receptor system at super‐resolution for natural killer and T‐cell biology

Killer‐cell immunoglobulin‐like receptors (KIRs) are components of two fundamental biological systems essential for human health and survival. First, they contribute to host immune responses, both innate and adaptive, through their expression by natural killer cells and T cells. Second, KIR play a key role in regulating placentation, and hence reproductive success. Analogous to the diversity of their human leucocyte antigen class I ligands, KIR are extremely polymorphic. In this review, we describe recent developments, fuelled by methodological advances, that are helping to decipher the KIR system in terms of haplotypes, polymorphisms, expression patterns and their ligand interactions. These developments are delivering deeper insight into the relevance of KIR in immune system function, evolution and disease.     

Putting the natural killer cell in its place

Natural killer (NK) cells were originally described as 'null' lymphocytes, but we have increasing evidence of their role in recognizing pathogen, and our knowledge of NK cell receptors continues to expand exponentially. Human NK cells have many receptors for human leucocyte antigen (HLA) class I. These killer immunoglobulin-like receptors (KIRs) and CD94/NKG2 receptors can signal in both positive and negative ways to regulate NK cell functions. The inhibitory receptors are the best characterized, but even in these cases much of their functional biology remains elusive. In this review, some recent advances in terms of the three-immunoglobulin (3Ig)-domain KIRs are discussed. Natural cytotoxicity receptors (NCRs) are among the activatory receptors found on NK cells. While pathogen ligands for these receptors have been described, endogenous ligands remain elusive. NCRs and NKG2D, a receptor for stress-induced antigens, appear to play complementary functional roles in terms of NK cell activation. More recently described on NK cells are the Toll-like receptors. In particular, these receptors of the innate immune system allow NK cells to directly sense pathogen, and their ligation on accessory cells indirectly activates NK cells through cytokine production. It is becoming clear that none of these receptor systems functions in isolation and that it is the sum of the signals (which will reflect the pathogenic situation), in addition to the cytokine milieu, that will direct NK cell activation. The resulting cytotoxicity, cytokine production and direct cell-cell regulatory interactions with other cells of the immune system, for example dendritic cells, ultimately determine the role of the NK cell in the context of an overall immune response.     

Evidence that the KIR2DS5 gene codes for a surface receptor triggering natural killer cell function

In this study, after immunization with NK cells from a KIR2DS5(+) donor and screening on cell transfectants expressing different members of the killer immunoglobulin-like receptor (KIR) family, we generated a mAb, DF200, reacting with several KIR2D receptors including KIR2DL1/L2/L3, KIR2DS1/S2 and KIR2DS5. By the analysis of peripheral blood NK cells and in vitro derived NK cell clones, we have demonstrated for the first time that KIR2DS5 is expressed at the cell surface in discrete subsets of NK cells and, after DF200 mAb-mediated engagement, can induce both cytotoxicity and cytokine release. Using co-transfection and co-immunoprecipitation, we found that KIR2DS5 associates with the DAP12 signaling polypeptide. Finally, soluble KIR2DS5-Fc fusion protein does not bind to cell transfectants expressing different HLA-C alleles, suggesting that, if KIR2DS5 does recognize HLA-C molecules, this may only occur in the presence of certain peptides.     

Increased number and function of natural killer cells in human immunodeficiency virus 1‐positive subjects co‐infected with herpes simplex virus 2

Natural killer (NK) cells bridge the interface between innate and adaptive immunity and are implicated in the control of herpes simplex virus 2 (HSV‐2) infection. In subjects infected with human immunodeficiency virus 1 (HIV‐1), the critical impact of the innate immune response on disease progression has recently come into focus. Higher numbers of NK cells are associated with lower HIV‐1 plasma viraemia. Individuals with the compound genotype of killer cell immunoglobulin‐like receptor (KIR) 3DS1 and human leucocyte antigen (HLA)‐Bw4‐80I, or who have alleles of KIR3DL1 that encode proteins highly expressed on the NK cell surface, have a significant delay in disease progression. We studied the effect of HSV‐2 co‐infection in HIV‐1‐infected subjects, and show that HSV‐2 co‐infection results in a pan‐lymphocytosis, with elevated absolute numbers of CD4⁺ and CD8⁺ T cells, and NK cells. The NK cells in HSV‐2 co‐infected subjects functioned more efficiently, with an increase in degranulation after in vitro stimulation. The number of NK cells expressing the activating receptors NKp30 and NKp46, and expressing KIR3DL1 or KIR3DS1, was inversely correlated with HIV‐1 plasma viral load in subjects mono‐infected with HIV‐1, but not in subjects co‐infected with HSV‐2. This suggests that HSV‐2 infection mediates changes within the NK cell population that may affect immunity in HIV‐1 infection.     

Impaired natural killer cell‐induced antibody‐dependent cell‐mediated cytotoxicity is associated with human immunodeficiency virus‐1 disease progression

This study evaluates the correlation between natural killer (NK) cell function and human immunodeficiency virus (HIV)-1 disease progression in 133 untreated HIV-1 positive Chinese subjects, including 41 former plasma donors (FPDs) and 92 men who have sex with men, and 35 HIV-negative controls. Flow cytometry was used to determine the abundance of NK cell subsets, the expression levels of receptor species, human leucocyte antigen (HLA) genotyping and the antibody-dependent cell-mediated cytotoxicity (ADCC) responses of NK cells. We observed a decreased expression of CD56^dimCD16⁺ NK cell subsets and an increased expression of CD56⁻CD16⁺ with HIV-1 infection. As well, the expression of activating and inhibitory receptors increased significantly in NK cells, but CD16 receptor levels and the NKG2A/NKG2C ratio were down-regulated with HIV-1 infection. ADCC responses were higher in elite controllers than in all other groups, and were correlated inversely with HIV-1 viral load but correlated positively with CD4 count only in FPDs. Furthermore, individuals infected for < 1 year have lower ADCC responses than those infected for > 1 year. We also observed a negative association between ADCC responses and viral load in those who carry the HLA-A*30/B*13/Cw*06 haplotype. The positive correlation between CD16 expression and ADCC responses and a negative correlation trend between CD158a and ADCC responses were also observed (P = 0·058). Our results showed that the ADCC response is associated with patients'' disease status, receptor expression levels, infection time and specific HLA alleles, which indicates that ADCC may offer protective effects against HIV-1 infection.     

Expression of inhibitory-motif killer immunoglobulin-like receptor, KIR2DL1, is increased in natural killer cells from women with pelvic endometriosis

Problem:  We investigated inhibitory and activation motif expression of killer immunoglobulin-like receptor (KIR) by natural killer (NK) cells, which may be pathogenetically involved in endometriosis.
Method of study:  We compared cells from 24 Japanese women laparoscopically diagnosed with endometriosis, to cells from 25 women with other laparoscopic diagnoses. KIR expression by NK cells was assessed in peripheral blood (PB) and peritoneal fluid (PF) by flow cytometry. Intracellular immunoreceptor tyrosine-based (IT) inhibitory and activation motifs (ITIM and ITAM) of KIR in PB was assessed by Western blotting.
Results:  ITIM-KIR expression by PB NK cells was significantly and similarly greater than ITAM-KIR expression in women with and without endometriosis. Percentages of CD56⁺ NK cells in PB and PF did not differ significantly between women with and without endometriosis; however, the percentage of CD158a⁺ cells among CD56⁺ NK cells in PB and PF was significantly higher in women with than without endometriosis.
Conclusions:  ITIM-KIR expressing NK cells might confer tolerance to peritoneal endometriotic implants.     

Activating and inhibitory killer immunoglobulin‐like receptors (KIR) in haploidentical haemopoietic stem cell transplantation to cure high‐risk leukaemias

A number of experimental studies have shown that natural killer (NK) cells can eliminate cancer cells and the mechanisms involved in this effect have been uncovered during the last two decades. Clinical data from haploidentical haematopoietic stem cell transplantation (haplo‐HSCT) revealed that NK cells were responsible for remarkably favourable effects in both adult and paediatric high‐risk leukaemias. NK receptors specific for major histocompatibility complex (MHC) class I molecules, including killer immunoglobulin (Ig)‐like receptors (KIR) and CD94/NKG2A, play a major role in the anti‐leukaemia effect (mediating either inhibitory or activating signals). Haplo‐ HSCT requires a heavy conditioning regimen for the patient and the use of large numbers of T cell‐depleted HSC to be grafted. After transplantation, natural killer cells develop from HSC shortly after engraftment and may include ‘alloreactive’ NK cells that kill leukaemic cells and prevent graft‐versus‐host disease (GvHD). Alloreactive NK cells are characterized by the expression of KIR that are not engaged by any of the human leucocyte antigen (HLA) class I alleles expressed by the patient. Their generation is dependent upon the existence of a KIR/HLA class I mismatch between donor and recipient. Novel important information on the function and specificity of different KIR has been obtained recently by the analysis of donor‐derived alloreactive NK cells in a cohort of paediatric patients given haplo‐HSCT to cure acute, high‐risk leukaemias.     

The extensive polymorphism of KIR genes

The functions of human natural killer (NK) cells are controlled by diverse families of antigen receptors. Prominent among these are the killer cell immunoglobulin‐like receptors (KIR), a family of genes clustered in one of the most variable regions of the human genome. Within this review we discuss the vast polymorphism of the KIR gene complex which rivals that of the human leucocyte antigen (HLA) complex. There are several aspects to this polymorphism. Initially there is presence/absence of individual KIR genes, with four of these genes, termed framework genes, being present in all individuals tested to date, except on those very occasional instances when the gene has been deleted. Within each gene, alleles are present at different frequencies. We provide details of a new website that enables convenient searching for data on KIR gene, allele and genotype frequencies in different populations and show how these frequencies vary in different worldwide populations and the high probability of individuals differing in their KIR repertoire when both gene and allele polymorphism is considered. The KIR genes present in an individual may be classified into A and/or B haplotypes, which respectively have a more inhibitory role or a more activating role on the function of the NK cell. Family studies have been used to ascertain the make‐up of these haplotypes, inclusion of allele typing enabling determination of whether one or two copies of a particular gene is present. In addition to genetic diversification the KIR gene complex shows differences at the functional level with different alleles having different protein expression levels and different avidity with their HLA ligand.     

KIR2DL3 and KIR2DL1 show similar impact on licensing of human NK cells

Killer cell immunoglobulin‐like receptor/HLA class I (KIR/HLA‐I) combinations are associated with disease risk, implicating functional roles for NK cells (NKCs) or KIR⁺ T cells. KIR/HLA‐I interactions can act through inhibition of NKC activation by target cells and NKC licensing for greater intrinsic responsiveness. We compared licensing conferred by the weaker, HLA‐C group 1/KIR2DL3, and the stronger, HLA‐C group 2/KIR2DL1, inhibitory combinations. The “rheostat model” predicts weaker licensing by HLA‐C1/KIR2DL3 interactions than HLA‐C2/KIR2DL1. We analyzed degranulation in NKC subsets expressing single and multiple receptors for HLA‐I. NKG2A had the strongest licensing impact, while KIR2DL3, KIR2DL1, and KIR3DL1 were weaker, and not significantly different to each other. Presence of one or two matched HLA‐C allotypes did not alter licensing of KIR2DL3⁺ and KIR2DL1⁺ NKC. Coexpression of activating KIR2DS1 disarmed KIR2DL3⁺ and KIR2DL1⁺ NKC to a similar extent. KIR3DL1 and NKG2A combined for more enhanced licensing of double‐positive NKC than the combination of KIR2DL3 and KIR2DL1. Thus, KIR2DL3 and KIR2DL1 have similar capacity to license NKC, suggesting that inhibitory signal strength and amount of available HLA‐C ligands do not correlate with NKC licensing. Altogether, our results show that the basis for disease associations of HLA‐C and KIR2DL likely encompasses factors other than licensing.     

Control of early stages in invariant natural killer T‐cell development

Natural killer T (NKT) cells develop in the thymus from the same precursors as conventional CD4(+) and CD8(+) αβ T cells, CD4(+) CD8(+) double-positive cells. In contrast to conventional αβT cells, which are selected by MHC-peptide complexes presented by thymic epithelial cells, invariant NKT cells are selected by lipid antigens presented by the non-polymorphic, MHC I-like molecule CD1d, present on the surface of other double-positive thymocytes, and require additional signals from the signalling lymphocytic-activation molecule (SLAM) family of receptors. In this review, we provide a discussion of recent findings that have modified our understanding of the NKT cell developmental programme, with an emphasis on events that affect the early stages of this process. This includes factors that control double-positive thymocyte lifespan, and therefore the ability to generate the canonical Vα rearrangements that characterize this lineage, as well as the signal transduction pathways engaged downstream of the T-cell receptor and SLAM molecules.     

Functional activity of natural killer cells and killer T cells in mice after ovarian transplantation

Tashkent Postgraduate Medical Institute, Ministry of Health of the USSR. (Presented by Academician of the Academy of Medical Sciences of the USSR N. A. Lopatkin). Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 112, No. 9, pp. 273–275, September, 1991.     

Influence of KIR gene diversity on the course of HSV-1 infection: resistance to the disease is associated with the absence of KIR2DL2 and KIR2DS2

Herpes simplex virus type 1 (HSV-1) causes lifelong latent infections in most humans. Periodical virus reactivations from latency in the neurons of sensitive ganglia lead to transport to mucocutaneous regions and productive replication, which results in recurrent inflammatory herpetic lesions or in asymptomatic virus shedding. The medical consequences of such lesions and the frequency of recurrences vary greatly in different subjects. Furthermore, many infected individuals never suffer manifestations of the disease, even when exposed to stimuli that trigger clinical recurrences in other humans. The origin of the variability in the clinical course of HSV-1 infection remains unexplained. Herpesviruses and other pathogens sabotage the expression of major histocompatibility complex class I molecules by infected cells, thus subverting T-cell-mediated immunity. Subversion of antigen presentation is counteracted by natural killer cells, which survey the human leukocyte antigen (HLA) expression by specific receptors. These include the killer cell immunoglobulin-like receptors (KIRs), which are encoded by a complex of extremely diverse and rapidly evolving genes. Here, we analyze the contribution of KIR gene diversity to the variable clinical course of HSV-1 infection by comparing the distribution of these genes in humans with clinical manifestations of the disease with that in asymptomatically infected donors. This study provides preliminary evidence that the receptors KIR2DL2 and KIR2DS2 predispose to symptomatic HSV-1 infection and favor the frequently recurring forms of the disease. Possible contribution of the 'HLA-C1' ligand to HSV-1 disease was not statistically supported. Because of an absolute genetic linkage between KIR2DL2 and KIR2DS2, we could not determine which receptor was primarily responsible for the observed association, but our results suggest that presence in the genome of KIR2DL2 and KIR2DS2 hinders an effective cellular response to HSV-1.