HLA-C cell surface expression and control of HIV/AIDS correlate with a variant upstream of HLA-C

Abstract

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.

Figure 1

Level of mRNA expression varies among HLA-C alleles and correlates with the -35 SNP. HLA-C was amplified from the cDNA of peripheral blood lymphocytes and the relative contribution of each allele was established by cloning and restriction digestion of PCR products. In donors heterozygous at the -35 genotype, HLA-C alleles in LD with the -35C genotype are in excess compared to the HLA-C allele that is in LD with -35T. The numbers of clones identified were as follows: Donor 1, Cw*0602 (-35C) 43 clones and Cw*0401 (-35T) 17 clones; Donor 2, Cw*0202 (-35C) 49 clones and Cw*0701 (-35T) 10 clones; Donor 3, Cw*0602 (-35C) 46 clones and Cw*0701 (-35T) 12 clones; Donor 4, Cw*0303 (-35T ) 20 clones and Cw*0701 (-35T) 29 clones. Statistical analysis was performed using two-tailed t-test with binomial distribution.

Figure 2

Variation in the level of surface HLA-C protein correlates with -35 genotype. HLA-C molecules on CD3⁺ T lymphocytes from 50 normal individuals were measured by flow cytometry using the DT9 mAb and grouped according to -35 genotype (a) and HLA-C allele (b). The -35 allele (C or T) that is in LD with the corresponding HLA-C allele is shown below each HLA-C allele name along the x axis and the -35 genotype of each donor is indicated by shading of the individual data points.

Figure 3

Highly expressed HLA-C allotypes control HIV to a greater extent than HLA C allotypes with lower expression. Odds ratios are based on the frequency of HLA-C alleles in the VL <2,000 group versus the >10,000 group. -35 LD with the respective HLA-C allele is indicated on the x axis.

Figure 4

The phylogram shows neighbor-joining relationships among full-length HLA-C alleles estimated using Kimura’s 2-parameter substitution model. Numbers above nodes indicate bootstrap recovery of that node in 500 bootstrap replications and the scale bar indicates the number of substitutions per site in a given interval along branches. HLA-C alleles known to be in strong LD with C at −35kb are indicated with a solid triangle while the two functional supertypes containing either Asn77 or Ser77 are distinguished by a ‘-N’ suffix on the designation of alleles containing an Asn residue. Strong bootstrap support (up to 100%) indicates that alleles associated with either C or T at -35 group together in multiple clades. Likewise, alleles of both functional supertypes are grouped together in multiple clades (bootstrap support up to 98%).