Natural killer (NK) cells provide a central defense against viral infection by using inhibitory and activation receptors for major histocompatibility complex class I molecules as a means of controlling their activity. We show that genes encoding the inhibitory NK cell receptor KIR2DL3 and its human leukocyte antigen C group 1 (HLA-C1) ligand directly influence resolution of hepatitis C virus (HCV) infection. This effect was observed in Caucasians and African Americans with expected low infectious doses of HCV but not in those with high-dose exposure, in whom the innate immune response is likely overwhelmed. The data strongly suggest that inhibitory NK cell interactions are important in determining antiviral immunity and that diminished inhibitory responses confer protection against HCV.

Natural killer (NK) cells provide defense in the early stages of the innate immune response against viral infections by producing cytokines and causing cytotoxicity. The killer immunoglobulin-like receptors (KIRs) on NK cells regulate the inhibition and activation of NK-cell responses through recognition of human leukocyte antigen (HLA) class I molecules on target cells KIR and HLA loci are both highly polymorphic, and some HLA class I products bind and trigger cell-surface receptors specified by KIR genes. Here we report that the activating KIR allele KIR3DS1, in combination with HLA-B alleles that encode molecules with isoleucine at position 80 (HLA-B Bw4-80Ile), is associated with delayed progression to AIDS in individuals infected with human immunodeficiency virus type 1 (HIV-1). In the absence of KIR3DS1, the HLA-B Bw4-80Ile allele was not associated with any of the AIDS outcomes measured. By contrast, in the absence of HLA-B Bw4-80Ile alleles, KIR3DS1 was significantly associated with more rapid progression to AIDS. These observations are strongly suggestive of a model involving an epistatic interaction between the two loci. The strongest synergistic effect of these loci was on progression to depletion of CD4(+) T cells, which suggests that a protective response of NK cells involving KIR3DS1 and its HLA class I ligands begins soon after HIV-1 infection.

The proteins encoded by the classical HLA class I and class II genes in the major histocompatibility complex (MHC) are highly polymorphic and play an essential role in self/nonself immune recognition. HLA variation is a crucial determinant of transplant rejection and susceptibility to a large number of infectious and autoimmune disease1. Yet identification of causal variants is problematic due to linkage disequilibrium (LD) that extends across multiple HLA and non-HLA genes in the MHC2,3. We therefore set out to characterize the LD patterns between the highly polymorphic HLA genes and background variation by typing the classical HLA genes and >7,500 common single nucleotide polymorphisms (SNPs) and deletion/insertion polymorphisms (DIPs) across four population samples. The analysis provides informative tag SNPs that capture some of the variation in the MHC region and that could be used in initial Corresponding author: John D. Rioux, Montreal Heart Institute, 5000 Rue Bélanger, Montréal, Québec, Canada, H1T 1C8, E-mail: rioux@broad.mit.edu. 16 These authors contributed equally All data will be available at the following sites: http://www.broad.mit.edu/mpg/idrg/projects/hla/ http://www.sanger.ac.uk/HGP/Chr6/ http://www.glovar.org COMPETING FINANCIAL INTEREST STATEMENTThe authors declare that they have no competing financial interests. Numerous studies have demonstrated association between HLA alleles and disease susceptibility (a partial list is provided in Table 1 and Supplementary Table 1), but the interpretation of these results is confounded by the strong correlation between alleles at neighboring HLA and non-HLA genes. Major efforts have therefore been directed at cataloguing the gene and variation content of the entire MHC4-6. In addition, previous studies in European-derived populations have examined the distribution of LD across the region and have suggested that SNPs could help dissect causal variation within the MHC2,3,7-10. Here, we have created a resource to guide future association studies by genotyping genetic variants across the extended MHC region of 7.5 Mb at a higher density and in more DNA samples than previously reported. In 361 individuals of African (YRI), European (CEU), Chinese (CHB), and Japanese (JPT) ancestry, the inferred haplotype structure across the region shows that LD is systematically higher in CEU, CHB and JPT samples than in the YRI sample (Fig. 1). Alleles across the different classical HLA loci demonstrate strong correlation (Supplementary Table 2). These high levels of LD among SNPs and DIPs and among HLA alleles suggest that SNPs outside the HLA genes are informative about HLA types (Fig. 2a), and that a few, well chosen SNPs may capture common classical HLA variation at several loci. Europe PMC Funders GroupWe examined the association between HLA types and single SNPs across the entire region. Fig. 2b shows the results for HLA-C (see Supplementary Fig. 1 for the other HLA genes). In the four populations studied, 34-44% of the HLA alleles present are strongly associa...

Allotypes of the natural killer (NK) cell receptor KIR3DL1 vary in both NK cell expression patterns and inhibitory capacity upon binding to their ligands, HLA-B Bw4 molecules, present on target cells. Using a sample size of over 1,500 human immunodeficiency virus (HIV)+ individuals, we show that various distinct allelic combinations of the KIR3DL1 and HLA-B loci significantly and strongly influence both AIDS progression and plasma HIV RNA abundance in a consistent manner. These genetic data correlate very well with previously defined functional differences that distinguish KIR3DL1 allotypes. The various epistatic effects observed here for common, distinct KIR3DL1 and HLA-B Bw4 combinations are unprecedented with regard to any pair of genetic loci in human disease, and indicate that NK cells may have a critical role in the natural history of HIV infection.

A variant upstream of human leukocyte antigen C (HLA-C) shows the most significant genome-wide effect on HIV control in European Americans and is also associated with the level of HLA-C expression. We characterized the differential cell surface expression levels of all common HLA-C allotypes and tested directly for effects of HLA-C expression on outcomes of HIV infection in 5243 individuals. Increasing HLA-C expression was associated with protection against multiple outcomes independently of individual HLA allelic effects in both African and European Americans, regardless of their distinct HLA-C frequencies and linkage relationships with HLA-B and HLA-A. Higher HLA-C expression was correlated with increased likelihood of cytotoxic T lymphocyte responses and frequency of viral escape mutation. In contrast, high HLA-C expression had a deleterious effect in Crohn’s disease, suggesting a broader influence of HLA expression levels in human disease.

This analysis shows that, in patients with HIV-1 infection, a single amino acid change in HLA molecules has a substantial effect on the rate of progression to AIDS. The different consequences of HLA-B*35-PY and HLA-B*35-Px in terms of disease progression highlight the importance of the epitope specificities of closely related class I molecules in the immune defense against HIV-1.

The HLA-C locus is distinct relative to the other classical HLA class I loci in that it has relatively limited polymorphism1, lower expression on the cell surface2,3, and more extensive ligand-receptor interactions with killer cell immunoglobulin-like receptors (KIR)4. A single nucleotide polymorphism (SNP) 35Kb upstream of HLA-C (rs9264942; termed −35) associates with control of HIV5–7, and with levels of HLA-C mRNA transcripts8 and cell surface expression7, but the mechanism underlying its varied expression is unknown. We proposed that the −35 SNP is not the causal variant for differential HLA-C expression, but rather is marking another polymorphism that directly affects levels of HLA-C7. Here we show that variation within the 3′ untranslated region of HLA-C regulates binding of the microRNA Hsa-miR-148a to its target site, resulting in relatively low surface expression of alleles that bind this microRNA and high expression of HLA-C alleles that escape post-transcriptional regulation. The 3′UTR variant associates strongly with control of HIV, potentially adding to the effects of genetic variation encoding the peptide-binding region of the HLA class I loci. Variation in HLA-C expression adds another layer of diversity to this highly polymorphic locus that must be considered when deciphering the function of these molecules in health and disease.

A variant 35 kb upstream of the HLA-C gene (-35C/T) was previously shown to associate with HLA-C mRNA expression level and steady-state plasma HIV RNA levels. We genotyped this variant in 1,698 patients of European ancestry with HIV. Individuals with known seroconversion dates were used for disease progression analysis and those with longitudinal viral load data were used for viral load analysis. We further tested cell surface expression of HLA-C in normal donors using an HLA-C-specific antibody. We show that the -35C allele is a proxy for high HLA-C cell surface expression, and that individuals with high-expressing HLA-C alleles progress more slowly to AIDS and control viremia significantly better than individuals with low HLA-C expressing alleles. These data strongly implicate high HLA-C expression levels in more effective control of HIV-1, potentially through better antigen presentation to cytotoxic T lymphocytes or recognition and killing of infected cells by natural killer cells.Correspondence should be addressed to M.C. (carringm@mail.nih.gov). Note: Supplementary information is available on the Nature Genetics website. Reprints and permissions information is available online at http://npg.nature.com/reprintsandpermissions/. AUTHOR CONTRIBUTIONS NIH Public Access Author ManuscriptNat Genet. Author manuscript; available in PMC 2010 December 1. Recently, however, a scan for genetic variants that influence the control of viral load indicated that a dimorphism 35 kb upstream of the HLA-C gene (-35C/T) had one of the two strongest genome-wide effects on the level of plasma viremia in early, established HIV infection as measured by viral load set-point, although no significant association of this variant was observed with progression to a CD4 cell count of <350 (ref.2 ). Notably, the -35C variant that associates with low viral load has also been shown to associate with high HLA-C mRNA levels in a codominant manner among a group of individuals of European ancestry 12,13 , although whether it associates with cell surface expression has not been tested. These findings suggest that certain HLA-C allotypes might have a primary role in restricting HIV replication through innate and/or acquired immune mechanisms that have previously been overlooked. Here we present data from 1,698 European American individuals, indicating that high levels of HLA-C confer strong protection early in the course of HIV infection and this early protection of high HLA-C extends to some extent into chronic infection. We propose a model in which highexpression HLA-C alleles might confer better innate and/or acquired immune responses than low-expression HLA-C alleles. RESULTS Effect of-35 on mean viral loadThe effect of -35 genotypes on mean plasma HIV load (mVL) measurements was tested in a group of 935 seroincident European American individuals (see Online Methods). Each individual was categorized into one of three groups based on their mVL (<2,000, 2,000-10,000 and >10,000 mean viral RNA copies per ml plasma), and the frequency of ea...