The success of the immune response is finely balanced between, on the one hand, the need to engage vigorously with, and clear, certain pathogens; and, on the other, the requirement to minimize immunopathology and autoimmunity. Distinct immune strategies to achieve this balance have evolved in females and males and also in infancy through to adulthood. Sex differences in outcome from a range of infectious diseases can be identified from as early as fetal life, such as in congenital cytomegalovirus infection. The impact of sex hormones on the T-helper 1/T-helper 2 cytokine balance has been proposed to explain the higher severity of most infectious diseases in males. In the minority where greater morbidity and mortality is observed in females, this is hypothesized to arise because of greater immunopathology and/or autoimmunity. However, a number of unexplained exceptions to this rule are described. Studies that have actually measured the sex differences in children in the immune responses to infectious diseases and that would further test these hypotheses, are relatively scarce.
sex; gender; pediatric; infections
The potential contribution of HLA-A alleles to viremic control in chronic HIV type 1 (HIV-1) infection has been relatively understudied compared with HLA-B. In these studies, we show that HLA-A*7401 is associated with favorable viremic control in extended southern African cohorts of >2100 C-clade–infected subjects. We present evidence that HLA-A*7401 operates an effect that is independent of HLA-B*5703, with which it is in linkage disequilibrium in some populations, to mediate lowered viremia. We describe a novel statistical approach to detecting additive effects between class I alleles in control of HIV-1 disease, highlighting improved viremic control in subjects with HLA-A*7401 combined with HLA-B*57. In common with HLA-B alleles that are associated with effective control of viremia, HLA-A*7401 presents highly targeted epitopes in several proteins, including Gag, Pol, Rev, and Nef, of which the Gag epitopes appear immunodominant. We identify eight novel putative HLA-A*7401–restricted epitopes, of which three have been defined to the optimal epitope. In common with HLA-B alleles linked with slow progression, viremic control through an HLA-A*7401–restricted response appears to be associated with the selection of escape mutants within Gag epitopes that reduce viral replicative capacity. These studies highlight the potentially important contribution of an HLA-A allele to immune control of HIV infection, which may have been concealed by a stronger effect mediated by an HLA-B allele with which it is in linkage disequilibrium. In addition, these studies identify a factor contributing to different HIV disease outcomes in individuals expressing HLA-B*5703.
There is progressive concern about the evolving burden of morbidity and mortality caused by coinfection with HIV-1 and hepatitis B virus (HBV) in sub-Saharan Africa, but the epidemiology and impact of this problem are not well defined. We therefore set out to assimilate more information about the nature of HBV/HIV coinfection in this region by undertaking a retrospective observational study of southern African adult women. We used samples from previously recruited HIV-1 positive women attending antenatal clinics in three settings in South Africa and Botswana (n = 950) and added a small cohort of HIV-negative antenatal South African women for comparison (n = 72). We tested for HBsAg and followed up HBsAg-positive samples by testing for HBeAg, HBV DNA, HBV genotype, presence of drug-resistance associated mutations (RAMs) and HDV. We identified HBsAg in 72 individuals (7% of the whole cohort), of whom 27% were HBeAg-positive, and the majority HBV genotypes A1 and A2. We did not detect any HDV coinfection. HBV prevalence was significantly different between geographically distinct cohorts, but did not differ according to HIV status. Among adults from South Africa, HBV/HIV coinfected patients had lower CD4+ T cell counts compared to those with HIV-monoinfection (p = 0.02), but this finding was not replicated in the cohort from Botswana. Overall, these data provide a snapshot of the coinfection problem at the heart of the HIV/HBV co-epidemic, and are important to inform public health policy, resource allocation, education, surveillance and clinical care.
HLA class I polymorphism has a major influence on adult HIV disease progression. An important mechanism mediating this effect is the impact on viral replicative capacity (VRC) of the escape mutations selected in response to HLA-restricted CD8+ T-cell responses. Factors that contribute to slow progression in pediatric HIV infection are less well understood. We here investigate the relationship between VRC and disease progression in pediatric infection, and the effect of HLA on VRC and on disease outcome in adult and pediatric infection. Studying a South African cohort of >350 ART-naïve, HIV-infected children and their mothers, we first observed that pediatric disease progression is significantly correlated with VRC. As expected, VRCs in mother-child pairs were strongly correlated (p = 0.004). The impact of the protective HLA alleles, HLA-B*57, HLA-B*58:01 and HLA-B*81:01, resulted in significantly lower VRCs in adults (p<0.0001), but not in children. Similarly, in adults, but not in children, VRCs were significantly higher in subjects expressing the disease-susceptible alleles HLA-B*18:01/45:01/58:02 (p = 0.007). Irrespective of the subject, VRCs were strongly correlated with the number of Gag CD8+ T-cell escape mutants driven by HLA-B*57/58:01/81:01 present in each virus (p = 0.0002). In contrast to the impact of VRC common to progression in adults and children, the HLA effects on disease outcome, that are substantial in adults, are small and statistically insignificant in infected children. These data further highlight the important role that VRC plays both in adult and pediatric progression, and demonstrate that HLA-independent factors, yet to be fully defined, are predominantly responsible for pediatric non-progression.
HLA plays a central role in determining disease outcome in adult HIV infection. A principal mechanism by which this HLA effect is mediated is via viral replicative capacity (VRC), protective HLA alleles such as HLA-B*57 driving the selection of viral escape mutants that reduce VRC. The factors contributing to the diverse disease progression rates observed in infected children, however, remain uncertain. We here address the role of HLA and VRC in pediatric disease progression in a large cohort in Kimberley, South Africa. The findings highlight the consistent and important role of VRC in both adult and pediatric progression. However, the impact of key HLA molecules in shaping disease outcome in adult infection is notably absent in pediatric infection. Further studies of pediatric infection therefore provide the potential to gain critical new insights into HLA-independent mechanisms of HIV disease non-progression that predominate in HIV-infected but healthy, ART-naive children. Understanding these mechanisms remains of direct relevance to the development of future interventions to minimize HIV disease.
parvovirus; PARV4; HIV; South Africa; prevalence; co-infection; serology; human; viruses
Mutations within cytotoxic T lymphocyte (CTL) epitopes impair T cell recognition, but escape mutations arising in flanking regions that alter antigen processing have not been defined in natural human infections. In human histocompatibility leukocyte antigen (HLA)-B57+ HIV-infected persons, immune selection pressure leads to a mutation from alanine to proline at Gag residue 146 immediately preceding the NH2 terminus of a dominant HLA-B57–restricted epitope, ISPRTLNAW. Although N-extended wild-type or mutant peptides remained well-recognized, mutant virus–infected CD4 T cells failed to be recognized by the same CTL clones. The A146P mutation prevented NH2-terminal trimming of the optimal epitope by the endoplasmic reticulum aminopeptidase I. These results demonstrate that allele-associated sequence variation within the flanking region of CTL epitopes can alter antigen processing. Identifying such mutations is of major relevance in the construction of vaccine sequences.
CD8 T cell responses; viral evolution; immune evasion; antigen presentation
Successful vaccine development for infectious diseases has largely been achieved in settings where natural immunity to the pathogen results in clearance in at least some individuals. HIV presents an additional challenge in that natural clearance of infection does not occur, and the correlates of immune protection are still uncertain. However, partial control of viremia and markedly different outcomes of disease are observed in HIV infected persons. Here we examine the antiviral mechanisms implicated by one variable that has been consistently associated with extremes of outcome, namely HLA class I alleles, and in particular HLA-B, and examine the mechanisms by which this modulation is likely to occur, and the impact of these interactions on evolution of the virus and the host. Studies to date provide evidence for both HLA-dependent and epitope-dependent influences on viral control and viral evolution, and have important implications for the continued quest for an effective HIV vaccine.
The strongest genetic influence on immune control in HIV-1 infection is the HLA class I genotype. Rapid disease progression in B-clade infection has been linked to HLA-B*35 expression, in particular to the less common HLA-B*3502 and HLA-B*3503 subtypes but also to the most prevalent subtype, HLA-B*3501. In these studies we first demonstrated that whereas HLA-B*3501 is associated with a high viral set point in two further B-clade-infected cohorts, in Japan and Mexico, this association does not hold in two large C-clade-infected African cohorts. We tested the hypothesis that clade-specific differences in HLA associations with disease outcomes may be related to distinct targeting of critical CD8+ T-cell epitopes. We observed that only one epitope was significantly targeted differentially, namely, the Gag-specific epitope NPPIPVGDIY (NY10, Gag positions 253 to 262) (P = 2 × 10−5). In common with two other HLA-B*3501-restricted epitopes, in Gag and Nef, that were not targeted differentially, a response toward NY10 was associated with a significantly lower viral set point. Nonimmunogenicity of NY10 in B-clade-infected subjects derives from the Gag-D260E polymorphism present in ∼90% of B-clade sequences, which critically reduces recognition of the Gag NY10 epitope. These data suggest that in spite of any inherent HLA-linked T-cell receptor repertoire differences that may exist, maximizing the breadth of the Gag-specific CD8+ T-cell response, by the addition of even a single epitope, may be of overriding importance in achieving immune control of HIV infection. This distinction is of direct relevance to development of vaccines designed to optimize the anti-HIV CD8+ T-cell response in all individuals, irrespective of HLA type.
Interleukin-10 (IL-10) is an immunoregulatory cytokine that influences the clinical outcome of chronic viral infections. Results show that IL-10-promoter genetic variants influence human immunodeficiency virus-1 pathogenesis possibly via regulating IL-10 plasma levels and the breadth of CD8+ T-cell immune responses.
Background. Interleukin-10 (IL-10) is a potent immunoregulatory cytokine. IL-10-promoter polymorphisms have been shown to affect human immunodeficiency virus type 1 (HIV-1) clinical outcomes but the underlying mechanisms are poorly understood.
Methods. We investigated the relationship between IL-10-promoter variants, plasma cytokine levels, immune responses and markers of disease outcome in antiretroviral-naïve HIV-1 chronically infected individuals from South Africa. Two IL-10-promoter single nucleotide polymorphisms (SNPs) were genotyped in 451 participants. Baseline plasma levels of select cytokines were measured for 112 individuals. Viral load, CD4+ T-cell counts and HIV-1-specific interferon-gamma CD8+ T-cell immune responses were measured at baseline. CD4+ T-cell counts were measured longitudinally and rates of CD4+ T-cell decline computed for 300 study subjects.
Results. The minor IL-10-1082G and -592A variants occurred at frequencies of 0.31 and 0.34, respectively. The -592AA genotype associated significantly with attenuated loss of CD4+ T cells (P = .0496). Individuals possessing -1082GG had significantly higher IL-10 levels compared to -1082AA/AG (P = .0006). The -592AA genotype was associated with greater breadth of virus-specific CD8+ T-cell responses compared to CC and CA (P = .002 and .004 respectively).
Conclusions. IL-10-promoter variants may influence the rate of HIV-1 disease progression by regulating IL-10 levels and the breadth of CD8+ T-cell immune responses.
HLA class I genotype is a major determinant of the outcome of HIV infection, and the impact of certain alleles on HIV disease outcome is well studied. Recent studies have demonstrated that certain HLA class I alleles that are in linkage disequilibrium, such as HLA-A*74 and HLA-B*57, appear to function co-operatively to result in greater immune control of HIV than mediated by either single allele alone. We here investigate the extent to which HLA alleles - irrespective of linkage disequilibrium - function co-operatively.
We here refined a computational approach to the analysis of >2000 subjects infected with C-clade HIV first to discern the individual effect of each allele on disease control, and second to identify pairs of alleles that mediate ‘co-operative additive’ effects, either to improve disease suppression or to contribute to immunological failure. We identified six pairs of HLA class I alleles that have a co-operative additive effect in mediating HIV disease control and four hazardous pairs of alleles that, occurring together, are predictive of worse disease outcomes (q<0.05 in each case). We developed a novel ‘sharing score’ to quantify the breadth of CD8+ T cell responses made by pairs of HLA alleles across the HIV proteome, and used this to demonstrate that successful viraemic suppression correlates with breadth of unique CD8+ T cell responses (p = 0.03).
These results identify co-operative effects between HLA Class I alleles in the control of HIV-1 in an extended Southern African cohort, and underline complementarity and breadth of the CD8+ T cell targeting as one potential mechanism for this effect.
HIV-1 attenuation resulting from immune escape mutations selected in Gag may contribute to slower disease progression in HIV-1-infected individuals expressing certain HLA class I alleles. We previously showed that the protective allele HLA-B*81 and the HLA-B*81-selected Gag T186S mutation are strongly associated with a lower viral replication capacity of recombinant viruses encoding Gag-protease derived from individuals chronically infected with HIV-1 subtype C. In the present study, we directly tested the effect of this mutation on viral replication capacity. In addition, we investigated potential compensatory effects of various polymorphisms, including other HLA-B*81-associated mutations that significantly covary with the T186S mutation. Mutations were introduced into a reference subtype B backbone and into patient-derived subtype C sequences in subtype B and C backbones by site-directed mutagenesis. The exponential-phase growth of mutant and wild-type viruses was assayed by flow cytometry of a green fluorescent protein reporter T cell line or by measurement of HIV-1 reverse transcriptase activity in culture supernatants. Engineering of the T186S mutation alone into all patient-derived subtype C sequences failed to yield replication-competent viruses, while in the subtype B sequence, the T186S mutation resulted in impaired replication capacity. Only the T186S mutation in combination with the T190I mutation yielded replication-competent viruses for all virus backbones tested; however, these constructs replicated slower than the wild type, suggesting that only partial compensation is mediated by the T190I mutation. Constructs encoding the T186S mutation in combination with other putative compensatory mutations were attenuated or defective. These results suggest that the T186S mutation is deleterious to HIV-1 subtype C replication and likely requires complex compensatory pathways, which may contribute to the clinical benefit associated with HLA-B*81.
Pediatric HIV-1 infection is characterized by rapid disease progression and without antiretroviral therapy (ART), more than 50% of infected children die by the age of 2 years. However, a small subset of infected children progresses slowly to disease in the absence of ART. This study aimed to identify functional characteristics of HIV-1-specific T cell responses that distinguish children with rapid and slow disease progression. Fifteen perinatally HIV-infected children (eight rapid and seven slow progressors) were longitudinally studied to monitor T cell polyfunctionality. HIV-1-specific interferon (IFN)-γ+ CD8+ T cell responses gradually increased over time but did not differ between slow and rapid progressors. However, polyfunctional HIV-1-specific CD8+ T cell responses, as assessed by the expression of four functions (IFN-γ, CD107a, TNF-α, MIP-1β), were higher in slow compared to rapid progressors (p=0.05) early in infection, and was associated with slower subsequent disease progression. These data suggest that the quality of the HIV-specific CD8+ T cell response is associated with the control of disease in children as has been shown in adult infection.
It is unknown whether favorable HLA class II alleles may attenuate HIV-1 through selection pressure in a manner similar to that of protective HLA class I alleles. We investigated the relationship between HLA class II alleles and in vitro replication capacities of recombinant viruses encoding HIV-1 subtype C Gag-protease from chronically infected individuals. No associations were found between individual alleles and lower replication capacity, suggesting no significant HIV-1 attenuation by HLA class II-restricted Gag-specific CD4+ T cell immune pressure.
Recent studies suggest that natural killer T (NKT) cells play a role in early antiviral pathogenesis and are rapidly depleted in chronic human immunodeficiency virus type 1 (HIV-1) clade B infection. We aimed to characterize the phenotypic and functional characteristics of NKT cells in HIV-1 clade C-infected Africans at different stages of HIV-1 disease. NKT cell frequencies, subsets, and ex vivo effector functions were assessed using multiparametric flow cytometry in a cross-sectional analysis of cryopreserved peripheral blood mononuclear cells from a cohort of 53 HIV-1 clade C chronically infected South African adults with CD4 T cell counts ranging from 94 to 839 cells/μl. We observed a significant decline of NKT cell numbers in advanced HIV-1 disease as well as activation and functional impairment of NKT cells in individuals with low CD4 T cell counts. The loss of NKT cells was largely driven by a reduction in the CD4+ and CD4–CD8– NKT cell subsets in advanced disease. These findings demonstrate significant impairment of the NKT cell compartment in progressive HIV-1 clade C disease that might play an important role in the modulation of immune function in HIV-1 infection.
The promiscuous presentation of epitopes by similar HLA class I alleles holds promise for a universal T-cell-based HIV-1 vaccine. However, in some instances, cytotoxic T lymphocytes (CTL) restricted by HLA alleles with similar or identical binding motifs are known to target epitopes at different frequencies, with different functional avidities and with different apparent clinical outcomes. Such differences may be illuminated by the association of similar HLA alleles with distinctive escape pathways. Using a novel computational method featuring phylogenetically corrected odds ratios, we systematically analyzed differential patterns of immune escape across all optimally defined epitopes in Gag, Pol, and Nef in 2,126 HIV-1 clade C-infected adults. Overall, we identified 301 polymorphisms in 90 epitopes associated with HLA alleles belonging to shared supertypes. We detected differential escape in 37 of 38 epitopes restricted by more than one allele, which included 278 instances of differential escape at the polymorphism level. The majority (66 to 97%) of these resulted from the selection of unique HLA-specific polymorphisms rather than differential epitope targeting rates, as confirmed by gamma interferon (IFN-γ) enzyme-linked immunosorbent spot assay (ELISPOT) data. Discordant associations between HLA alleles and viral load were frequently observed between allele pairs that selected for differential escape. Furthermore, the total number of associated polymorphisms strongly correlated with average viral load. These studies confirm that differential escape is a widespread phenomenon and may be the norm when two alleles present the same epitope. Given the clinical correlates of immune escape, such heterogeneity suggests that certain epitopes will lead to discordant outcomes if applied universally in a vaccine.
HLA class I-mediated selection of immune escape mutations in functionally important Gag epitopes may partly explain slower disease progression in HIV-1-infected individuals with protective HLA alleles. To investigate the impact of Gag function on disease progression, the replication capacities of viruses encoding Gag-protease from 60 individuals in early HIV-1 subtype C infection were assayed in an HIV-1-inducible green fluorescent protein reporter cell line and were correlated with subsequent disease progression. Replication capacities did not correlate with viral load set points (P = 0.37) but were significantly lower in individuals with below-median viral load set points (P = 0.03), and there was a trend of correlation between lower replication capacities and lower rates of CD4 decline (P = 0.09). Overall, the proportion of host HLA-specific Gag polymorphisms in or adjacent to epitopes was negatively associated with replication capacities (P = 0.04), but host HLA-B-specific polymorphisms were associated with higher viral load set points (P = 0.01). Further, polymorphisms associated with host-specific protective HLA alleles were linked with higher viral load set points (P = 0.03). These data suggest that transmission or early HLA-driven selection of Gag polymorphisms results in reduced early cytotoxic T-lymphocyte (CTL) responses and higher viral load set points. In support of the former, 46% of individuals with nonprotective alleles harbored a Gag polymorphism exclusively associated with a protective HLA allele, indicating a high rate of their transmission in sub-Saharan Africa. Overall, HIV disease progression is likely to be affected by the ability to mount effective Gag CTL responses as well as the replication capacity of the transmitted virus.
One proposed HIV vaccine strategy is to induce Gag-specific CD8+ T-cell responses that can corner the virus, through fitness cost of viral escape and unavailability of compensatory mutations. We show here that the most variable capsid residues principally comprise escape mutants driven by protective alleles HLA-B*57, -5801, and -8101 and covarying HLA-independent polymorphisms that arise in conjunction with these escape mutations. These covarying polymorphisms are potentially compensatory and are concentrated around three tropism-determining loops of p24, suggesting structural interdependencies. Our results demonstrate complex patterns of adaptation of HIV under immune selection pressure, the understanding of which should aid vaccine design.
The association between HLA-B*2705 and the immune control of human immunodeficiency virus type 1 (HIV-1) has previously been linked to the targeting of the HLA-B*2705-restricted Gag epitope KRWIILGLNK (KK10) by CD8+ T cells. In order to better define the mechanisms of the HLA-B*2705 immune control of HIV, we first characterized the CD8+ T-cell responses of nine highly active antiretroviral therapy (HAART)-naïve B*2705-positive subjects. Unexpectedly, we observed a strong response to an HLA-B*2705-restricted Pol epitope, KRKGGIGGY (KY9), in 8/9 subjects. The magnitude of the KY9 response was only marginally lower than that of the KK10-specific response (median, 695 versus 867 spot-forming cells [SFC]/million peripheral blood mononuclear cells [PBMCs]; not significant [NS]), and viral escape mutants were observed in both KY9 and KK10, resulting from selection pressure driven by the respective CD8+ T-cell response. By comparing inhibitions of viral replication by CD8+ T cells specific for the Gag KK10, Pol KY9, and Vpr VL9 HLA-B*2705-restricted epitopes, we observed a consistent hierarchy of antiviral efficacy (Gag KK10 > Pol KY9 > Vpr VL9). This hierarchy was associated with early recognition of HIV-1-infected cells, within 6 h of infection, by KK10- and KY9-specific CD8+ T cells but not until 18 h postinfection by VL9-specific CD8+ T cells. There was no association between antiviral efficacy and proliferative capacity, cytotoxicity, polyfunctionality, or T-cell receptor (TCR) avidity. These data are consistent with previous studies indicating an important role for the B*2705-Gag KK10 response in the control of HIV but also suggest a previously unrecognized role played by the subdominant Pol-specific KY9 response in HLA-B*2705-mediated control of HIV and that the recognition of HIV-infected cells by CD8+ T cells early in the viral life cycle may be important for viral containment in HIV-infected individuals.
The mechanisms underlying HIV-1 control by protective HLA class I alleles are not fully understood and could involve selection of escape mutations in functionally important Gag epitopes resulting in fitness costs. This study was undertaken to investigate, at the population level, the impact of HLA-mediated immune pressure in Gag on viral fitness and its influence on HIV-1 pathogenesis. Replication capacities of 406 recombinant viruses encoding plasma-derived Gag-protease from patients chronically infected with HIV-1 subtype C were assayed in an HIV-1-inducible green fluorescent protein reporter cell line. Viral replication capacities varied significantly with respect to the specific HLA-B alleles expressed by the patient, and protective HLA-B alleles, most notably HLA-B*81, were associated with lower replication capacities. HLA-associated mutations at low-entropy sites, especially the HLA-B*81-associated 186S mutation in the TL9 epitope, were associated with lower replication capacities. Most mutations linked to alterations in replication capacity in the conserved p24 region decreased replication capacity, while most in the highly variable p17 region increased replication capacity. Replication capacity also correlated positively with baseline viral load and negatively with baseline CD4 count but did not correlate with the subsequent rate of CD4 decline. In conclusion, there is evidence that protective HLA alleles, in particular HLA-B*81, significantly influence Gag-protease function by driving sequence changes in Gag and that conserved regions of Gag should be included in a vaccine aiming to drive HIV-1 toward a less fit state. However, the long-term clinical benefit of immune-driven fitness costs is uncertain given the lack of correlation with longitudinal markers of disease progression.
HIV can be partially contained by host immunity and understanding the basis of this may inform vaccine design. The importance of B-cell function in long-term control is poorly understood. One method of investigating this is in vivo cellular depletion. In this study, we take advantage of a unique opportunity to investigate the role of B cells in an HIV-infected patient. The HIV-1+ patient studied here was not taking antiretroviral drugs and was treated for pre-existing low-grade lymphoplasmacytoid lymphoma by depletion of CD20+ B cells using rituximab. We demonstrate that B-cell depletion results in a decline in autologous neutralizing antibody (NAb) responses and a 1.7 log10 rise in HIV-1 plasma viral load (pVL). The recovery of NAbs results in a decline in pVL. The HIV-1 sequences diversify and NAb-resistant mutants are subsequently selected. These data suggest that B-cell function can contribute to the long-term control of pVL, and that NAbs may be more important in controlling chronic HIV-1 infection than previously suspected.
HIV infection can be partially regulated by the host immune system; however whether B cells contribute to this response is unclear. Huang et al. show that transient depletion of B cells can result in an increase in HIV viral load suggesting that these immune cells do participate in the control of HIV infection.
The recent failure of the T-cell-based HIV vaccine trial led by Merck & Co., Inc. prompts the urgent need to refocus on the question of which T-cell responses are required to control HIV replication. The well-described association between the expression of particular MHC class I molecules and successful containment of HIV or, in the macaque model, SIV replication provide a valuable starting point from which to evaluate more precisely what might constitute effective CD8+ T-cell responses. Here, we review recent studies of T-cell-mediated control of HIV and SIV infection, and offer insight for the design of a successful T-cell-based HIV vaccine in the future.
HIV-1 specific HLA-B-restricted CD8+ T cell responses differ from HLA-C-restricted responses in antiviral effectiveness. To investigate possible reasons for these differences, we characterized the frequency and polyfunctionality of immmunodominant HLA-B*57/B5801- and HLA-Cw*07-restricted CD8+ T cells occurring concurrently in nine study subjects assessing IFN-γ, TNF-α, IL-2, MIP-1β, and CD107a by flow cytometry and analyzed sequence variation in targeted epitopes. HLA-B*57/5801 and HLA-Cw*07 restricted CD8+ T cells did not differ significantly in polyfunctionality (p = 0.84). Possession of three or more functions correlated positively with CD4+ T cell counts (r = 0.85; p = 0.006) and monofunctional CD8+ T cells inversely correlated with CD4 cell counts (r = −0.79; p = 0.05). There were no differences in polyfunctionality of CD8+ T cells specific to wildtype versus mutated epitopes. These results suggest that loss of polyfunctionality and increase in monofunctional HIV-1-specific CD8+ T cells are associated with disease progression independent of restricting HLA allele. Furthermore, sequence variation does not appear to significantly impact CD8+ T cell polyfunctionality in chronic HIV-1 infection.
HLA-B*57/5801; HLA-C; HIV-1 chronic infection; CD8+ T cells; Polyfunctionality
Despite high potential for HIV-1 genetic variation, the emergence of some mutations is constrained by fitness costs, and may be associated with compensatory amino acid (AA) co-variation. To characterize the interplay between Cytotoxic T Lymphocyte (CTL)-mediated pressure and HIV-1 evolutionary pathways, we investigated AA co-variation in Gag sequences obtained from 449 South African individuals chronically infected with HIV-1 subtype C.
Individuals with CTL responses biased toward Gag presented lower viral loads than individuals with under-represented Gag-specific CTL responses. Using methods that account for founder effects and HLA linkage disequilibrium, we identified 35 AA sites under Human Leukocyte Antigen (HLA)-restricted CTL selection pressure and 534 AA-to-AA interactions. Analysis of two-dimensional distances between co-varying residues revealed local stabilization mechanisms since 40% of associations involved neighboring residues. Key features of our co-variation analysis included sites with a high number of co-varying partners, such as HLA-associated sites, which had on average 55% more connections than other co-varying sites.
Clusters of co-varying AA around HLA-associated sites (especially at typically conserved sites) suggested that cooperative interactions act to preserve the local structural stability and protein function when CTL escape mutations occur. These results expose HLA-imprinted HIV-1 polymorphisms and their interlinked mutational paths in Gag that are likely due to opposite selective pressures from host CTL-mediated responses and viral fitness constraints.