A commonly recognized simian immunodeficiency virus Nef epitope presented to cytotoxic T lymphocytes of Indian-origin rhesus monkeys by the prevalent major histocompatibility complex class I allele Mamu-A*02

The ability to monitor vaccine-elicited CD8(+) cytotoxic T-lymphocyte (CTL) responses in simian immunodeficiency virus (SIV)- and simian-human immunodeficiency virus (SHIV)-infected rhesus monkeys has been limited by our knowledge of viral epitopes predictably presented to those lymphocytes by common rhesus monkey MHC class I alleles. We now define an SIV and SHIV Nef CTL epitope (YTSGPGIRY) that is presented to CD8(+) T lymphocytes by the common rhesus monkey MHC class I molecule Mamu-A*02. All seven infected Mamu-A*02(+) monkeys evaluated demonstrated this response, and peptide-stimulated interferon gamma Elispot assays indicated that the response represents a large proportion of the entire CD8(+) T-lymphocyte SIV- or SHIV-specific immune response of these animals. Knowledge of this epitope and MHC class I allele substantially increases the number of available rhesus monkeys that can be used for testing prototype HIV vaccines in this important animal model.     

Mamu-A*01 allele-mediated attenuation of disease progression in simian-human immunodeficiency virus infection

Expression of several major histocompatibility complex (MHC) class I alleles is associated with a protective effect against disease progression in both human immunodeficiency virus type 1 and simian immunodeficiency virus infection. To understand the mechanism underlying this effect, we investigated the expression of the MHC class I allele Mamu-A*01 in simian-human immunodeficiency virus (SHIV) infection, one of the major models for evaluation of AIDS vaccine candidates. We found that disease progression was significantly delayed in Mamu-A*01-positive rhesus monkeys infected with the highly pathogenic SHIV 89.6P. The delay corresponded not only to a noted Mamu-A*01-restricted dominant cytotoxic T-lymphocyte (CTL) response but also to a lower viral load in lymph nodes (LN) and, importantly, to minimal destruction of LN structure during early infection. In contrast, Mamu-A*01-negative monkeys exhibited massive destruction of LN structure with accompanying rapid disease progression. These data indicate that MHC class I allele-restricted CTL responses may play an important role in preservation of lymphoid tissue structure, thereby resulting in attenuation of disease progression in immunodeficiency virus infection.     

Vaccine-elicited memory cytotoxic T lymphocytes contribute to Mamu-A*01-associated control of simian/human immunodeficiency virus 89.6P replication in rhesus monkeys

The expression of particular major histocompatibility complex (MHC) class I alleles can influence the rate of disease progression following lentiviral infections. This effect is a presumed consequence of potent cytotoxic T-lymphocyte (CTL) responses that are restricted by these MHC class I molecules. The present studies have examined the impact of the MHC class I allele Mamu-A*01 on simian/human immunodeficiency virus 89.6P (SHIV-89.6P) infection in unvaccinated and vaccinated rhesus monkeys by exploring the contribution of dominant-epitope specific CTL in this setting. Expression of Mamu-A*01 in immunologically naive monkeys was not associated with improved control of viral replication, CD4+ T-lymphocyte loss, or survival. In contrast, Mamu-A*01+ monkeys that had received heterologous prime/boost immunizations prior to challenge maintained higher CD4+ T-lymphocyte levels and better control of SHIV-89.6P replication than Mamu-A*01- monkeys. This protection was associated with the evolution of high-frequency anamnestic CTL responses specific for a dominant Mamu-A*01-restricted Gag epitope following infection. These data indicate that specific MHC class I alleles can confer protection in the setting of a pathogenic SHIV infection by their ability to elicit memory CTL following vaccination.     

Peptide-binding motifs associated with MHC molecules common in Chinese rhesus macaques are analogous to those of human HLA supertypes and include HLA-B27-like alleles

Chinese rhesus macaques are of particular interest in simian immunodeficiency virus/human immunodeficiency virus (SIV/HIV) research as these animals have prolonged kinetics of disease progression to acquired immunodeficiency syndrome (AIDS), compared to their Indian counterparts, suggesting that they may be a better model for HIV. Nevertheless, the specific mechanism(s) accounting for these kinetics remains unclear. The study of major histocompatibility complex (MHC) molecules, including their MHC/peptide-binding motifs, provides valuable information for measuring cellular immune responses and deciphering outcomes of infection and vaccine efficacy. In this study, we have provided detailed characterization of six prevalent Chinese rhesus macaque MHC class I alleles, yielding a combined phenotypic frequency of 29 %. The peptide-binding specificity of two of these alleles, Mamu-A2*01:02 and Mamu-B*010:01, as well as the previously characterized allele Mamu-B*003:01 (and Indian rhesus Mamu-B*003:01), was found to be analogous to that of alleles in the HLA-B27 supertype family. Specific alleles in the HLA-B27 supertype family, including HLA-B*27:05, have been associated with long-term nonprogression to AIDS in humans. All six alleles characterized in the present study were found to have specificities analogous to HLA supertype alleles. These data contribute to the concept that Chinese rhesus macaque MHC immunogenetics is more similar to HLA than their Indian rhesus macaque counterparts and thereby warrants further studies to decipher the role of these alleles in the context of SIV infection.     

CD8(+) lymphocytes from simian immunodeficiency virus-infected rhesus macaques recognize 14 different epitopes bound by the major histocompatibility complex class I molecule mamu-A*01: implications for vaccine design and testing

It is becoming increasingly clear that any human immunodeficiency virus (HIV) vaccine should induce a strong CD8(+) response. Additional desirable elements are multispecificity and a focus on conserved epitopes. The use of multiple conserved epitopes arranged in an artificial gene (or EpiGene) is a potential means to achieve these goals. To test this concept in a relevant disease model we sought to identify multiple simian immunodeficiency virus (SIV)-derived CD8(+) epitopes bound by a single nonhuman primate major histocompatibility complex (MHC) class I molecule. We had previously identified the peptide binding motif of Mamu-A*01(2), a common rhesus macaque MHC class I molecule that presents the immunodominant SIV gag-derived cytotoxic T lymphocyte (CTL) epitope Gag_CM9 (CTPYDINQM). Herein, we scanned SIV proteins for the presence of Mamu-A*01 motifs. The binding capacity of 221 motif-positive peptides was determined using purified Mamu-A*01 molecules. Thirty-seven peptides bound with apparent K(d) values of 500 nM or lower, with 21 peptides binding better than the Gag_CM9 peptide. Peripheral blood mononuclear cells from SIV-infected Mamu-A*01(+) macaques recognized 14 of these peptides in ELISPOT, CTL, or tetramer analyses. This study reveals an unprecedented complexity and diversity of anti-SIV CTL responses. Furthermore, it represents an important step toward the design of a multiepitope vaccine for SIV and HIV.     

Escape in one of two cytotoxic T-lymphocyte epitopes bound by a high-frequency major histocompatibility complex class I molecule,Mamu-A*02: a paradigm for virus evolution and persistence?

It is now accepted that an effective vaccine against AIDS must include effective cytotoxic-T-lymphocyte (CTL) responses. The simian immunodeficiency virus (SIV)-infected rhesus macaque is the best available animal model for AIDS, but analysis of macaque CTL responses has hitherto focused mainly on epitopes bound by a single major histocompatibility complex (MHC) class I molecule, Mamu-A*01. The availability of Mamu-A*01-positive macaques for vaccine studies is therefore severely limited. Furthermore, it is becoming clear that different CTL responses are able to control immunodeficiency virus replication with varying success, making it a priority to identify and analyze CTL responses restricted by common MHC class I molecules other than Mamu-A*01. Here we describe two novel epitopes derived from SIV, one from Gag (Gag(71-79) GY9), and one from the Nef protein (Nef(159-167) YY9). Both epitopes are bound by the common macaque MHC class I molecule, Mamu-A*02. The sequences of these two eptiopes are consistent with the molecule's peptide-binding motif, which we have defined by elution of natural ligands from Mamu-A*02. Strikingly, we found evidence for the selection of escape variant viruses by CTL specific for Nef(159-167) YY9 in 6 of 6 Mamu-A*02-positive animals. In contrast, viral sequences encoding the Gag(71-79) GY9 epitope remained intact in each animal. This situation is reminiscent of Mamu-A*01-restricted CTL that recognize Tat(28-35) SL8, which reproducibly selects for escape variants during acute infection, and Gag(181-189) CM9, which does not. Differential selection by CTL may therefore be a paradigm of immunodeficiency virus infection.     

Major histocompatibility complex class I alleles associated with slow simian immunodeficiency virus disease progression bind epitopes recognized by dominant acute-phase cytotoxic-T-lymphocyte responses

Certain major histocompatibility complex class I (MHC-I) alleles are associated with delayed disease progression in individuals infected with human immunodeficiency virus (HIV) and in macaques infected with simian immunodeficiency virus (SIV). However, little is known about the influence of these MHC alleles on acute-phase cellular immune responses. Here we follow 51 animals infected with SIV(mac)239 and demonstrate a dramatic association between Mamu-A*01 and -B*17 expression and slowed disease progression. We show that the dominant acute-phase cytotoxic T lymphocyte (CTL) responses in animals expressing these alleles are largely directed against two epitopes restricted by Mamu-A*01 and one epitope restricted by Mamu-B*17. One Mamu-A*01-restricted response (Tat(28-35)SL8) and the Mamu-B*17-restricted response (Nef(165-173)IW9) typically select for viral escape variants in early SIV(mac)239 infection. Interestingly, animals expressing Mamu-A*1 and -B*17 have less variation in the Tat(28-35)SL8 epitope during chronic infection than animals that express only Mamu-A*01. Our results show that MHC-I alleles that are associated with slow progression to AIDS bind epitopes recognized by dominant CTL responses during acute infection and underscore the importance of understanding CTL responses during primary HIV infection.     

Conserved MHC class I peptide binding motif between humans and rhesus macaques

Since the onset of the HIV pandemic, the use of nonhuman primate models of infection has increasingly become important. An excellent model to study HIV infection and immunological responses, in particular cell-mediated immune responses, is SIV infection of rhesus macaques. CTL epitopes have been mapped using SIV-infected rhesus macaques, but, to date, a peptide binding motif has been described for only one rhesus class I MHC molecule, Mamu-A*01. Herein, we have established peptide-live cell binding assays for four rhesus MHC class I molecules: Mamu-A*11, -B*03, -B*04, and -B*17. Using such assays, peptide binding motifs have been established for all four of these rhesus MHC class I molecules. With respect to the nature and spacing of crucial anchor positions, the motifs defined for Mamu-B*04 and -B*17 present unique features not previously observed for other primate species. The motifs identified for Mamu-A*11 and -B*03 are very similar to the peptide binding motifs previously described for human HLA-B*44 and -B*27, respectively. Accordingly, naturally processed peptides derived from HLA-B*44 and HLA-B*27 specifically bind Mamu-A*11 and Mamu-B*03, respectively, indicating that conserved MHC class I binding capabilities exist between rhesus macaques and humans. The definition of four rhesus MHC class I-specific motifs expands our ability to accurately detect and quantitate immune responses to MHC class I-restricted epitopes in rhesus macaques and to rationally design peptide epitope-based model vaccine constructs destined for use in nonhuman primates.     

Identification of seventeen new simian immunodeficiency virus-derived CD8+ T cell epitopes restricted by the high frequency molecule, Mamu-A*02, and potential escape from CTL recognition

MHC class I-restricted CD8+ T cells play an important role in controlling HIV and SIV replication. In SIV-infected Indian rhesus macaques (Macaca mulatta), comprehensive CD8+ T cell epitope identification has only been undertaken for two alleles, Mamu-A*01 and Mamu-B*17. As a result, these two molecules account for virtually all known MHC class I-restricted SIV-derived CD8+ T cell epitopes. SIV pathogenesis research and vaccine testing have intensified the demand for epitopes restricted by additional MHC class I alleles due to the shortage of Mamu-A*01+ animals. Mamu-A*02 is a high frequency allele present in over 20% of macaques. In this study, we characterized the peptide binding of Mamu-A*02 using a panel of single amino acid substitution analogues and a library of 497 unrelated peptides. Of 230 SIVmac239 peptides that fit the Mamu-A*02 peptide-binding motif, 75 peptides bound Mamu-A*02 with IC50 values of < or = 500 nM. We assessed the antigenicity of these 75 peptides using an IFN-gamma ELISPOT assay with freshly isolated PBMC from eight Mamu-A*02+ SIV-infected macaques and identified 17 new epitopes for Mamu-A*02. The synthesis of five Mamu-A*02 tetramers demonstrated the discrepancy between tetramer binding and IFN-gamma secretion by SIV-specific CD8+ T cells during chronic SIV infection. Bulk sequencing determined that 2 of the 17 epitopes accumulated amino acid replacements in SIV-infected macaques by the chronic phase of infection, suggestive of CD8+ T cell escape in vivo. This work enhances the use of the SIV-infected macaque model for HIV and increases our understanding of the breadth of CD8+ T cell responses in SIV infection.     

Diverse peptide presentation of rhesus macaque major histocompatibility complex class I Mamu-A 02 revealed by two peptide complex structures and insights into immune escape of simian immunodeficiency virus

Major histocompatibility complex class I (MHC I)-restricted CD8(+) T-cell responses play a pivotal role in anti-human immunodeficiency virus (HIV) immunity and the control of viremia. The rhesus macaque is an important animal model for HIV-related research. Among the MHC I alleles of the rhesus macaque, Mamu-A 02 is prevalent, presenting in ≥20% of macaques. In this study, we determined the crystal structure of Mamu-A 02, the second structure-determined MHC I from the rhesus macaque after Mamu-A 01. The peptide presentation characteristics of Mamu-A 02 are exhibited in complex structures with two typical Mamu-A 02-restricted CD8(+) T-cell epitopes, YY9 (Nef159 to -167; YTSGPGIRY) and GY9 (Gag71 to -79; GSENLKSLY), derived from simian immunodeficiency virus (SIV). These two peptides utilize similar primary anchor residues (Ser or Thr) at position 2 and Tyr at position 9. However, the central region of YY9 is different from that of GY9, a difference that may correlate with the immunogenic variance of these peptides. Further analysis indicated that the distinct conformations of these two peptides are modulated by four flexible residues in the Mamu-A 02 peptide-binding groove. The rare combination of these four residues in Mamu-A 02 leads to a variant presentation for peptides with different residues in their central regions. Additionally, in the two structures of the Mamu-A 02 complex, we compared the binding of rhesus and human β(2) microglobulin (β(2)m) to Mamu-A 02. We found that the peptide presentation of Mamu-A 02 is not affected by the interspecies interaction with human β(2)m. Our work broadens the understanding of CD8(+) T-cell-specific immunity against SIV in the rhesus macaque.     

Expression of the major histocompatibility complex class I molecule Mamu-A*01 is associated with control of simian immunodeficiency virus SIVmac239 replication

Several HLA alleles are associated with attenuated human immunodeficiency virus disease progression. We explored the relationship between the expression of particular major histocompatibility complex (MHC) class I alleles and viremia in simian immunodeficiency virus SIV(mac)239-infected macaques. Of the common MHC class I alleles, animals that expressed Mamu-A*01 exhibited the best control of viral replication.     

Initiation of antiretroviral therapy during chronic SIV infection leads to rapid reduction in viral loads and the level of T-cell immune response

In the present era of increasing resistance of human immunodeficiency virus (HIV) to antiviral drugs, exploration of adjunct therapies directed at immune responses in combination with antiretroviral drugs may be of value for the treatment of acquired immunodeficiency syndrome. In this study, we designed a model for immune therapy using SIVmac251 infection in rhesus macaques. We explored the outcomes of primary infection on viral loads and the resulting T-cell immune responses in primates. The SIV-infected rhesus macaque model exhibited features similar to those observed in HIV-1 infection of humans. Major histocompatibility complex (MHC) segregation with viral loads were found to associate with viral containment and hence the duration of the disease-free latency period. Thus a better understanding of the relative roles of MHC class I allele in control of viral replication may provide important information for prophylactic or therapeutic vaccine designs. Mamu-A01 is significantly associated with higher immune response and control of viral replication. This allele is frequent in rhesus macaques of Indian origin (22%). Interestingly, Mamu-B01 (26% animals) was associated with lower immune responses and higher viral loads. Another allele, A08 was also predominantly present in 37% of the animals in this study. We observed higher viral replication in individual SIV-infected rhesus monkeys that did not demonstrate strong cellular immune responses. The results are important for understanding SIV disease progression in different MHC Mamu alleles and also for improving the interpretation and quality of pre-clinical studies in rhesus monkeys.     

ALVAC-SIV-gag-pol-env-based vaccination and macaque major histocompatibility complex class I (A*01) delay simian immunodeficiency virus SIVmac-induced immunodeficiency.

T-cell-mediated immune effector mechanisms play an important role in the containment of human immunodeficiency virus/simian immunodeficiency virus (HIV/SIV) replication after infection. Both vaccination- and infection-induced T-cell responses are dependent on the host major histocompatibility complex classes I and II (MHC-I and MHC-II) antigens. Here we report that both inherent, host-dependent immune responses to SIVmac251 infection and vaccination-induced immune responses to viral antigens were able to reduce virus replication and/or CD4+ T-cell loss. Both the presence of the MHC-I Mamu-A*01 genotype and vaccination of rhesus macaques with ALVAC-SIV-gag-pol-env (ALVAC-SIV-gpe) contributed to the restriction of SIVmac251 replication during primary infection, preservation of CD4+ T cells, and delayed disease progression following intrarectal challenge exposure of the animals to SIV(mac251 (561)). ALVAC-SIV-gpe immunization induced cytotoxic T-lymphocyte (CTL) responses cumulatively in 67% of the immunized animals. Following viral challenge, a significant secondary virus-specific CD8+ T-cell response was observed in the vaccinated macaques. In the same immunized macaques, a decrease in virus load during primary infection (P = 0.0078) and protection from CD4 loss during both acute and chronic phases of infection (P = 0.0099 and P = 0.03, respectively) were observed. A trend for enhanced survival of the vaccinated macaques was also observed. Neither boosting the ALVAC-SIV-gpe with gp120 immunizations nor administering the vaccine by the combination of mucosal and systemic immunization routes increased significantly the protective effect of the ALVAC-SIV-gpe vaccine. While assessing the role of MHC-I Mamu-A*01 alone in the restriction of viremia following challenge of nonvaccinated animals with other SIV isolates, we observed that the virus load was not significantly lower in Mamu-A*01-positive macaques following intravenous challenge with either SIV(mac251 (561)) or SIV(SME660). However, a significant delay in CD4+ T-cell loss was observed in Mamu-A*01-positive macaques in each group. Of interest, in the case of intravenous or intrarectal challenge with the chimeric SIV/HIV strains SHIV(89.6P) or SHIV(KU2), respectively, MHC-I Mamu-A*01-positive macaques did not significantly restrict primary viremia. The finding of the protective effect of the Mamu-A*01 molecule parallels the protective effect of the B*5701 HLA allele in HIV-1-infected humans and needs to be accounted for in the evaluation of vaccine efficacy against SIV challenge models.     

Simian immunodeficiency virus evades a dominant epitope-specific cytotoxic T lymphocyte response through a mutation resulting in the accelerated dissociation of viral peptide and MHC class I

The ability of an AIDS virus to escape from immune containment by selective mutation away from recognition by CTL was explored in simian immunodeficiency virus of macaques (SIVmac)-infected rhesus monkeys. CTL recognition of a previously defined common viral mutation in an immunodominant SIVmac Gag epitope was evaluated. CTL were assessed for their ability to recognize a SIVmac Gag protein with a single residue 2 (T --> A) replacement in the minimal epitope peptide bound by the MHC class I molecule Mamu-A*01. SIVmac Gag-specific CTL lysed Mamu-A*01+ target cells infected with recombinant vaccinia virus expressing the wild-type but not the mutant Gag protein. In addition, CTL recognized the mutant epitope peptide less efficiently than the wild-type virus peptide. In studies to determine the mechanism by which the mutant virus evaded CTL recognition, this peptide was shown to bind Mamu-A*01 in a manner that was indistinguishable from the wild-type peptide. However, experiments in which an increasing duration of delay was introduced between peptide sensitization of target cells and the assessment of these cells as targets in killing assays suggest that the mutant peptide with a T --> A replacement had a higher off-rate from Mamu-A*01 than the wild-type peptide did. Therefore, these findings suggest that AIDS viruses can evade virus-specific CTL responses through the accelerated dissociation of mutant peptide from MHC class I.     

Vectored Gag and Env but not Tat show efficacy against simian-human immunodeficiency virus 89.6P challenge in Mamu-A*01-negative rhesus monkeys

Simian-human immunodeficiency virus (SHIV) challenge studies in rhesus macaques were conducted to evaluate the efficacy of adenovirus-based vaccines in the context of different major histocompatibility complex class I genetic backgrounds and different vaccine compositions. Mamu-A*01 allele-negative rhesus monkeys were immunized with one of the following vaccine constructs: (i) replication-defective recombinant adenovirus type 5 (Ad5) expressing human immunodeficiency virus type 1 (HIV-1) Tat (Ad5/HIVTat); (ii) Ad5 vector expressing simian immunodeficiency virus (SIV) Gag (Ad5/SIVGag); (iii) Ad5 vector expressing the truncated HIV-1(jrfl) Env, gp140 (Ad5/gp140_jrfl); (iv) Ad5 vector expressing the SHIV-89.6P gp140 (Ad5/gp140_89.6P); or (v) the combination of Ad5/SIVGag and Ad5/gp140_jrfl. Following intravenous challenge with SHIV-89.6P, only those cohorts that received vaccines expressing Gag or Env exhibited an attenuation of the acute viremia and associated CD4-cell lymphopenia. While no prechallenge neutralizing antibody titers were detectable in either Ad5/gp140-vaccinated group, an accelerated neutralizing antibody response was observed in the Ad5/gp140_89.6P-vaccinated group upon viral challenge. The set-point viral loads in the Ad5/SIVGag- and Ad5/gp140_jrfl-vaccinated groups were associated with the overall strength of the induced cellular immune responses. To examine the contribution of Mamu-A*01 allele in vaccine efficacy against SHIV-89.6P challenge, Mamu-A*01-positive monkeys were immunized with Ad5/SIVGag. Vaccine-mediated protection was significantly more pronounced in the Mamu-A*01-positive monkeys than in Mamu-A*01-negative monkeys, suggesting the strong contributions of T-cell epitopes restricted by the Mamu-A*01 molecule. The implications of these results in the development of an HIV-1 vaccine will be discussed.     

Use of Major Histocompatibility Complex Class I/Peptide/β2M Tetramers To Quantitate CD8+ Cytotoxic T Lymphocytes Specific for Dominant and Nondominant Viral Epitopes in Simian-Human Immunodeficiency Virus-Infected Rhesus Monkeys

To evaluate the impact of the diversity of antigen recognition by T lymphocytes on disease pathogenesis, we must be able to identify and analyze simultaneously cytotoxic T-lymphocyte (CTL) responses specific for multiple viral epitopes. Many of the studies of the role of CD8(+) CTLs in AIDS pathogenesis have been done with simian immunodeficiency virus (SIV)- and simian-human immunodeficiency virus (SHIV)-infected rhesus monkeys. These studies have frequently made use of the well-defined SIV Gag CTL epitope p11C,C-M presented to CTL by the HLA-A homologue molecule Mamu-A*01. In the present study we identified and fine mapped two novel Mamu-A*01-restricted CTL epitopes: the SIVmac Pol-derived epitope p68A (STPPLVRLV) and the human immunodeficiency virus type 1 (HIV-1) Env-derived p41A epitope (YAPPISGQI). The frequency of CD8(+) CTLs specific for the p11C,C-M, p68A, and p41A epitopes was quantitated in the same animals with a panel of tetrameric Mamu-A*01/peptide/beta2m complexes. All SHIV-infected Mamu-A*01(+) rhesus monkeys tested had a high frequency of SIVmac Gag-specific CTLs to the p11C,C-M epitope. In contrast, only a fraction of the monkeys tested had detectable CTLs specific for the SIVmac Pol p68A and HIV-1 Env p41A epitopes, and these responses were detected at very low frequencies. Thus, the p11C,C-M-specific CD8(+) CTL response is dominant and the p41A- and p68A-specific CD8(+) CTL responses are nondominant. These results indicate that CD8(+) CTL responses to dominant CTL epitopes can be readily quantitated with the tetramer technology; however, CD8(+) CTL responses to nondominant epitopes, due to the low frequency of these epitope-specific cells, may be difficult to detect and quantitate by this approach.     

Immunodomination in the evolution of dominant epitope-specific CD8+ T lymphocyte responses in simian immunodeficiency virus-infected rhesus monkeys

Because the control of HIV-1 replication is largely dependent on CD8+ T lymphocyte responses specific for immunodominant viral epitopes, vaccine strategies that increase the breadth of dominant epitope-specific responses should contribute to containing HIV-1 spread. Developing strategies to elicit such broad immune responses will require an understanding of the mechanisms responsible for focusing CD8+ T lymphocyte recognition on a limited number of epitopes. To explore this biology, we identified cohorts of rhesus monkeys that expressed the MHC class I molecules Mamu-A*01, Mamu-A*02, or both, and assessed the evolution of their dominant epitope-specific CD8+ T lymphocyte responses (Gag p11C- and Tat TL8-specific in the Mamu-A*01+ and Nef p199RY-specific in the Mamu-A*02+ monkeys) following acute SIV infection. The Mamu-A*02+ monkeys that also expressed Mamu-A*01 exhibited a significant delay in the evolution of the CD8+ T lymphocyte responses specific for the dominant Mamu-A*02-restricted SIV epitope, Nef p199RY. This delay in kinetics was not due to differences in viral load kinetics or magnitude or in viral escape mutations, but was associated with the evolution of the Mamu-A*01-restricted CD8+ T lymphocyte responses to the highly dominant SIV epitopes Gag p11C and Tat TL8. Thus, the evolution of dominant epitope-specific CD8+ T lymphocyte responses can be suppressed by other dominant epitope-specific responses, and this immunodomination is important in determining the kinetics of dominant epitope-specific responses.     

The role of MHC class I allele Mamu-A*07 during SIVmac239 infection

Virus-specific CD8(+) T cells play an important role in controlling HIV/SIV replication. These T cells recognize intracellular pathogen-derived peptides displayed on the cell surface by individual MHC class I molecules. In the SIV-infected rhesus macaque model, five Mamu class I alleles have been thoroughly characterized with regard to peptide binding, and a sixth was shown to be uninvolved. In this study, we describe the peptide binding of Mamu-A1*007:01 (formerly Mamu-A*07), an allele present in roughly 5.08% of Indian-origin rhesus macaques (n?=?63 of 1,240). We determined a preliminary binding motif by eluting and sequencing endogenously bound ligands. Subsequently, we used a positional scanning combinatorial library and panels of single amino acid substitution analogs to further characterize peptide binding of this allele and derive a quantitative motif. Using this motif, we selected and tested 200 peptides derived from SIV(mac)239 for their capacity to bind Mamu-A1*007:01; 33 were found to bind with an affinity of 500?nM or better. We then used PBMC from SIV-infected or vaccinated but uninfected, A1*007:01-positive rhesus macaques in IFN-γ Elispot assays to screen the peptides for T-cell reactivity. In all, 11 of the peptides elicited IFN-γ(+) T-cell responses. Six represent novel A1*007:01-restricted epitopes. Furthermore, both Sanger and ultradeep pyrosequencing demonstrated the accumulation of amino acid substitutions within four of these six regions, suggestive of selective pressure on the virus by antigen-specific CD8(+) T cells. Thus, it appears that Mamu-A1*007:01 presents SIV-derived peptides to antigen-specific CD8(+) T cells and is part of the immune response to SIV(mac)239.     

The simian immunodeficiency virus envelope glycoprotein contains two epitopes presented by the Mamu-A*01 class I molecule.

Cytotoxic T lymphocyte (CTL) responses against the simian immunodeficiency virus (SIV) envelope and Gag proteins were monitored in a Mamu-A*01-positive rhesus macaque infected with SIVsmE660. Peripheral blood mononuclear cells (PBMC) cultured with synthetic peptides spanning the entire gp160 and Gag coding region recognized a total of three epitopes. One located in Gag was identified as the previously described Mamu-A*01-restricted p11cC-->M epitope (CTPYDINQM). The other two epitopes, designated p15m and p54m, were located in the gp160 envelope protein. Both were nine amino acids in length and were predicted to bind Mamu-A*01 because they contained proline and leucine residues at positions 3 and 9, respectively. Indeed, expression of this class I major histocompatibility complex molecule was required for target cell recognition by envelope-specific CD8(+) T cells directed against both epitopes. These Mamu-A*01-restricted epitopes in the SIV envelope will be useful for monitoring immune responses in vaccinated or infected animals.