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Portable flanking sequences modulate CTL epitope processing.

ABSTRACT: Peptide presentation is critical for immune recognition of pathogen-infected cells by CD8+ T lymphocytes. Although a limited number of immunodominant peptide epitopes are consistently observed in diseases such as HIV-1 infection, the relationship between immunodominance and antigen processing in humans is largely unknown. Here, we have demonstrated that endogenous processing and presentation of a human immunodominant HIV-1 epitope is more efficient than that of a subdominant epitope. Furthermore, we have shown that the regions flanking the immunodominant epitope constitute a portable motif that increases the production and antigenicity of otherwise subdominant epitopes. We used a novel in vitro degradation assay involving cytosolic extracts as well as endogenous intracellular processing assays to examine 2 well-characterized HIV-1 Gag overlapping epitopes presented by the same HLA class I allele, one of which is consistently immunodominant and the other subdominant in infected persons. The kinetics and products of degradation of HIV-1 Gag favored the production of peptides encompassing the immunodominant epitope and destruction of the subdominant one. Notably, cytosolic digestion experiments revealed flanking residues proximal to the immunodominant epitope that increased the production and antigenicity of otherwise subdominant epitopes. Furthermore, specific point mutations in these portable flanking sequences modulated the production and antigenicity of epitopes. Such portable epitope processing determinants provide what we believe is a novel approach to optimizing CTL responses elicited by vaccine vectors.

SUBMITTER: Le Gall S

PROVIDER: S-EPMC2045603 | biostudies-literature | 2007 Nov

REPOSITORIES: biostudies-literature

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Portable flanking sequences modulate CTL epitope processing.

Le Gall Sylvie S Stamegna Pamela P Walker Bruce D BD

The Journal of clinical investigation 20071101 11

Peptide presentation is critical for immune recognition of pathogen-infected cells by CD8+ T lymphocytes. Although a limited number of immunodominant peptide epitopes are consistently observed in diseases such as HIV-1 infection, the relationship between immunodominance and antigen processing in humans is largely unknown. Here, we have demonstrated that endogenous processing and presentation of a human immunodominant HIV-1 epitope is more efficient than that of a subdominant epitope. Furthermore ...[more]

PMID: 17975674

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Project description:Cytotoxic CD8+ T cells (CTLs) play a critical role in controlling viral infections. HIV-infected individuals develop CTL responses against epitopes derived from viral proteins, but also against cryptic epitopes encoded by viral alternative reading frames (ARF). We studied here the mechanisms of HIV-1 escape from CTLs targeting one such cryptic epitope, Q9VF, encoded by an HIVgag ARF and presented by HLA-B*07. Using PBMCs of HIV-infected patients, we first cloned and sequenced proviral DNA encoding for Q9VF. We identified several polymorphisms with a minority of proviruses encoding at position 5 an aspartic acid (Q9VF/5D) and a majority encoding an asparagine (Q9VF/5N). We compared the prevalence of each variant in PBMCs of HLA-B*07+ and HLA-B*07- patients. Proviruses encoding Q9VF/5D were significantly less represented in HLA-B*07+ than in HLA-B*07- patients, suggesting that Q9FV/5D encoding viruses might be under selective pressure in HLA-B*07+ individuals. We thus analyzed ex vivo CTL responses directed against Q9VF/5D and Q9VF/5N. Around 16% of HLA-B*07+ patients exhibited CTL responses targeting Q9VF epitopes. The frequency and the magnitude of CTL responses induced with Q9VF/5D or Q9VF/5N peptides were almost equal indicating a possible cross-reactivity of the same CTLs on the two peptides. We then dissected the cellular mechanisms involved in the presentation of Q9VF variants. As expected, cells infected with HIV strains encoding for Q9VF/5D were recognized by Q9VF/5D-specific CTLs. In contrast, Q9VF/5N-encoding strains were neither recognized by Q9VF/5N- nor by Q9VF/5D-specific CTLs. Using in vitro proteasomal digestions and MS/MS analysis, we demonstrate that the 5N variation introduces a strong proteasomal cleavage site within the epitope, leading to a dramatic reduction of Q9VF epitope production. Our results strongly suggest that HIV-1 escapes CTL surveillance by introducing mutations leading to HIV ARF-epitope destruction by proteasomes.

Project description:As human immunodeficiency virus (HIV) begins to replicate within hosts, immune responses are elicited against it. Escape mutations in viral epitopes-immunogenic peptide parts presented on the surface of infected cells-allow HIV to partially evade these responses, and thus rapidly go to fixation. The faster they go to fixation, i.e., the higher their escape rate, the larger the selective pressure exerted by the immune system is assumed to be. This relation underpins the rationale for using escapes to assess the strength of immune responses. However, escape rate estimates are often obtained by employing an aggregation procedure, where several mutations that affect the same epitope are aggregated into a single, composite epitope mutation. The aggregation procedure thus rests upon the assumption that all within-epitope mutations have indistinguishable effects on immune recognition. In this study, we investigate how violation of this assumption affects escape rate estimates. To this end, we extend a previously developed simulation model of HIV that accounts for mutation, selection, and recombination to include different distributions of fitness effects (DFEs) and inter-mutational genomic distances. We use this discrete time Wright-Fisher based model to simulate early within-host evolution of HIV for DFEs and apply standard estimation methods to infer the escape rates. We then compare true with estimated escape rate values. We also compare escape rate values obtained by applying the aggregation procedure with values estimated without use of that procedure. We find that across the DFEs analyzed, the aggregation procedure alters the detectability of escape mutations: large-effect mutations are overrepresented while small-effect mutations are concealed. The effect of the aggregation procedure is similar to extracting the largest-effect mutation appearing within an epitope. Furthermore, the more pronounced the over-exponential decay of the DFEs, the more severely true escape rates are underestimated. We conclude that the aggregation procedure has two main consequences. On the one hand, it leads to a misrepresentation of the DFE of fixed mutations. On the other hand, it conceals within-epitope interactions that may generate irregularities in mutation frequency trajectories that are thus left unexplained.

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Portable flanking sequences modulate CTL epitope processing.

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Portable flanking sequences modulate CTL epitope processing.

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