Identification and characterization of immunologically relevant CD8+ T cell epitopes of HIV-1 nef protein
HIV-1 mutates to escape HLA Class I restricted CD8+ T cells. Positively selected mutations (PSMs) are amino acid substitutions that allow the virus to survive under host immune pressure. However, the impact of PSMs on host clinical outcomes remains unknown. Negative effector (Nef), an accessory protein of HIV-1, enhances the pathogenicity and replication of the virus by down-regulating CD4 and HLA Class I expression from host cell surface to avoid recognition by immune cells, and modification of host cell signal transduction pathways to ensure a persistent state of infection. In both human and macaque models of infection some Nef-specific CD8+ T cell responses that drive viral mutations have been associated with better immune control. In this study, we identified PSMs within HIV-1 Nef, and analyzed their association with host immune selection pressure, and their potential impact on host disease status. We hypothesized that CD8+ T cell epitopes in HIV-1 Nef protein contain PSMs that are associated with different clinical outcomes in the infected host. HIV-1 nef gene was sequenced using 454 sequencing technology from 508 HIV-positive samples of the Pumwani sex-workers cohort of Nairobi, Kenya. PSMs in Nef were identified using bioinformatics tool, Quasi analysis and correlated with disease progression data. Three PSMs were associated with faster CD4 decline (E63D p=0.028, log rank: 4.799; I101V p=0.003, log rank:8.667 and I168M p=0.042, log rank:4.150), while two PSMs were associated with slower CD4 decline (H116N p=0.00011, log rank:14.891 and K182M p=0.03, log rank=4.753). Forty peptides containing a specific PSM were tested with ELISPOT assay, 27 of which were confirmed as CD8+ T cell epitopes. There was no significant difference in the frequency of antigen-specific CD8+ T cells with antiviral intracellular cytokines, their proliferation and exhaustion characteristics between peptides with PSMs associated with different disease progression status. However, the frequency of CD8+ T cells restricted by A*02:01-ILDLWVYNT (IT9-N) epitope, containing a PSM associated with slower CD4 decline, was higher (p0.0001) than CD8+ T cells restricted by A*02:01-ILDLWVYHT (IT9-H) epitope, containing consensus amino acid associated with faster CD4 decline. Identification of PSMs associated with different clinical outcomes and characterization of CD8+ T cell populations specific to these PSM containing epitopes can help to determine better immunogens for an effective HIV-1 vaccine.