Summary of the publications
Nozomi Kuse, Tomohiro Akahoshi, Hiroyuki Gatanaga, Takamasa Ueno, Shinichi Oka, and Masafumi Takiguchi, Selection of TI8-8V Mutant Associated with Long-Term Control of HIV-1 by Cross-Reactive HLA-B*51:01-restricted Cytotoxic T Cells, J. Immunol. 193:4814-4822, 2014

Elite controllers of HIV-1-infected HLA-B*51:01(+) hemophiliacs, who remain disease free and have a very low plasma viral load for >30 y, had the 8V mutation at an immunodominant Pol283-8 (TI8) epitope, whereas the 8T mutant was predominantly selected in other HIV-1-infected HLA-B*51:01(+) hemophiliacs, suggesting an important role of the 8V mutant selection in long-term control of HIV-1. However, the mechanism of this selection and the long-term control in these elite controllers remains unknown. In this study, we investigated the mechanism of the 8V mutant selection in these controllers. TI8-specific CTLs from these individuals evenly recognized both TI8 peptide-pulsed and TI8-8V peptide-pulsed cells and effectively suppressed replication of wild-type (WT) and the 8V viruses. However, the results of a competitive viral suppression assay demonstrated that CTLs from the individual who had WT virus could discriminate WT virus from the 8V virus, whereas those from the individuals who had the 8V virus evenly recognized both viruses. The former CTLs carried TCRs with weaker affinity for the HLA-B*51:01-TI8-8V molecule than for the HLA-B*51:01-TI-8 one, whereas the latter ones carried TCRs with similar affinity for both molecules. The reconstruction of the TCRs from these CTLs in TCR-deficient cells confirmed the different recognition of the TCRs for these epitopes. The present study showed that the 8V mutant virus could be selected by cross-reactive CTLs carrying TCR that could discriminate a small difference between the two molecules. The selection of the 8V mutant and elicitation of these two cross-reactive CTLs may contribute to the long-term control of HIV-1.

Xiaoming Sun*, Mamoru Fujiwara*, Yi Shi*, Nozomi Kuse, Hiroyuki Gatanaga, Victor Appay, George F. Gao, Shinichi Oka, and Masafumi Takiguchi (* Equal contribution), Superimposed epitopes restricted by the same HLA molecule drive distinct HIV-specific CD8+ T cell repertoires. J. Immunol. 193:77-84, 2014

Superimposed epitopes, in which a shorter epitope is embedded within a longer one, can be presented by the same HLA class I molecule. CD8(+) CTL responses against such epitopes and the contribution of this phenomenon to immune control are poorly characterized. In this study, we examined HLA-A*24:02-restricted CTLs specific for the superimposed HIV Nef epitopes RYPLTFGWCF (RF10) and RYPLTFGW (RW8). Unexpectedly, RF10-specific and RW8-specific CTLs from HIV-1-infected HLA-A*24:02+ individuals had no overlapping Ag reactivity or clonotypic compositions. Single-cell TCR sequence analyses demonstrated that RF10-specific T cells had a more diverse TCR repertoire than did RW8-specific T cells. Furthermore, RF10-specific CTLs presented a higher Ag sensitivity and HIV suppressive capacity compared with RW8-specific CTLs. Crystallographic analyses revealed important structural differences between RF10- and RW8-HLA-A*24:02 complexes as well, with featured and featureless conformations, respectively, providing an explanation for the induction of distinct T cell responses against these epitopes. The present study shows that a single viral sequence containing superimposed epitopes restricted by the same HLA molecule could elicit distinct CD8+ T cell responses, therefore enhancing the control of HIV replication. This study also showed that a featured epitope (e.g., RF10) could drive the induction of T cells with high TCR diversity and affinity.

Center for AIDS Research Best Paper Award 2014
Takayuki Chikata
, Jonathan M. Carlson, Yoshiko Tamura, Mohamed Ali Borghan, Takuya Naruto, Masao Hashimoto, Hayato Murakoshi, Anh Q. Le, Simon Mallal, Mina John, Hiroyuki Gatanaga, Shinichi Oka, Zabrina L. Brumme, and Masafumi Takiguchi, Host-specific adaptation of HIV-1 subtype B in the Japanese population, J.Virol. 88:4764-4775, 2014
The extent to which HIV-1 clade B strains exhibit population-specific adaptations to host HLA alleles remains incompletely known, in part due to incomplete characterization of HLA-associated HIV-1 polymorphisms (HLA-APs) in different global populations. Moreover, it remains unknown to what extent the same HLA alleles may drive significantly different escape pathways across populations. As the Japanese population exhibits distinctive HLA class I allele distributions, comparative analysis of HLA-APs between HIV-1 clade B-infected Japanese and non-Asian cohorts could shed light on these questions. However HLA-APs remain incompletely mapped in Japan. In a cohort of 430 treatment-naive Japanese with chronic HIV-1 clade B infection, we identified 284 HLA-APs in Gag, Pol and Nef using phylogenetically-corrected methods. The number of HLA-associated substitutions in Pol, notably those restricted by HLA-B*52:01, was weakly inversely correlated with plasma viral load (pVL), suggesting that the transmission and persistence of B*52:01-driven Pol mutations could modulate pVL. Differential selection of HLA-APs between HLA subtype members, including those differing only with respect to substitutions outside the peptide-binding groove, was observed, meriting further investigation as to their mechanisms of selection. Notably, two-thirds of HLA-AP identified in Japan had not been reported in previous studies of predominantly Caucasian cohorts, and were attributable to HLA alleles unique to, or enriched in, Japan. We also identified 71 cases where the same HLA allele drove significantly different escape pathways between Japan versus predominantly Caucasian cohorts. Our results underscore the distinct global evolution of HIV-1 clade B as a result of host population-specific cellular immune pressures.

IMPORTANCE SECTION: CTL escape mutations in HIV-1 are broadly predictable based on the HLA class I alleles expressed by the host. Because HLA allele distributions differ among worldwide populations, the pattern and diversity of HLA-associated escape mutations are likely to be somewhat distinct to each race and region. HLA-associated polymorphisms (HLA-APs) in HIV-1 have previously been identified at the population level in European, North American, Australian and African cohorts, however, large-scale analyses of HIV-1 clade B-specific HLA-APs in Asians are lacking. Differential intra-clade HIV-1 adaptation to global populations can be investigated via comparative analyses of HLA-associated polymorphisms across ethnic groups, but such studies are rare. Here, we identify HLA-APs in a large Japanese HIV-1 clade B cohort using phylogeneticaly-informed methods and observe that the majority of them had not been previously characterized in predominantly Caucasian populations. Results highlight HIV's unique adaptation to cellular immune pressures imposed by different global populations.

Chihiro Motozono*, Nozomi Kuse*, Xiaoming Sun, Pierre J. Rizkallah, Anna Fuller, Shinichi Oka, David K. Cole, Andrew K. Sewell,and Masafumi Takiguchi (*Equal contribution), Molecular basis of a dominant T-cell response to an HIV reverse transcriptase 8-mer epitope presented by the protective allele HLA-B*51:01, J. Immunol. 192:3428-3434, 2014
CD8(+) CTL responses directed toward the HLA-B*51:01-restricted HIV-RT128-135 epitope TAFTIPSI (TI8) are associated with long-term nonprogression to AIDS. Clonotypic analysis of responses to B51-TI8 revealed a public clonotype using TRAV17/TRBV7-3 TCR genes in six out of seven HLA-B*51:01(+) patients. Structural analysis of a TRAV17/TRBV7-3 TCR in complex with HLA-B51-TI8, to our knowledge the first human TCR complexed with an 8-mer peptide, explained this bias, as the unique combination of residues encoded by these genes was central to the interaction. The relatively featureless peptide-MHC (pMHC) was mainly recognized by the TCR CDR1 and CDR2 loops in an MHC-centric manner. A highly conserved residue Arg(97) in the CDR3 loop played a major role in recognition of peptide and MHC to form a stabilizing ball-and-socket interaction with the MHC and peptide, contributing to the selection of the public TCR clonotype. Surface plasmon resonance equilibrium binding analysis showed the low affinity of this public TCR is in accordance with the only other 8-mer interaction studied to date (murine 2C TCR-H-2K(b)-dEV8). Like pMHC class II complexes, 8-mer peptides do not protrude out the MHC class I binding groove like those of longer peptides. The accumulated evidence suggests that weak affinity might be a common characteristic of TCR binding to featureless pMHC landscapes.
Center for AIDS Research Best Paper Award 2013
Kristin Ladell*, Masao Hashimoto*, Maria Candela Iglesias*, Pascal G. Wilmann*, James E. McLaren, Stephanie Gras, Takayuki Chikata, Nozomi Kuse, Solene Fastenackels, Emma Gostick, John S. Bridgeman, Vanessa Venturi, Zaina Ait Arkoub, Henri Agut, David J. van Bockel, Jorge R. Almeida, Daniel C. Douek, Laurence Meyer, Alain Venet, Masafumi Takiguchi**, Jamie Rossjohn**, David A. Price** and Victor Appay** (*,** Equal contribution), A molecular basis for the control of preimmune escape variants by HIV-specific CD8+ T cells. Immunity. 38:425-436, 2013
The capacity of the immune system to adapt to rapidly evolving viruses is a primary feature of effective immunity, yet its molecular basis is unclear. Here, we investigated protective HIV-1-specific CD8+ T cell responses directed against the immunodominant p24 Gag-derived epitope KK10 (KRWIILGLNK263-272) presented by human leukocyte antigen (HLA)-B?2705. We found that cross-reactive CD8+ T cell clonotypes were mobilized to counter the rapid emergence of HIV-1 variants that can directly affect T cell receptor (TCR) recognition. These newly recruited clonotypes expressed TCRs that engaged wild-type and mutant KK10 antigens with similar affinities and almost identical docking modes, thereby accounting for their antiviral efficacy in HLA-B?2705+ individuals. A protective CD8+ T cell repertoire therefore encompasses the capacity to control TCR-accessible mutations, ultimately driving the development of more complex viral escape variants that disrupt antigen presentation.
Yuichi Yagita*, Nozomi Kuse*, Kimiko Kuroki*, Hiroyuki Gatanaga, Jonathan M. Carlson, Takayuki Chikata, Zabrina L. Brumme, Hayato Murakoshi, Tomohiro Akahoshi, Nico Pfeifer, Simon Mallal, Mina John, Toyoyuki Ose, Haruki Matsubara, Ryo Kanda, Yuko Fukunaga, Kazutaka Honda, Yuka Kawashima, Yasuo Ariumi, Shinichi Oka, Katsumi Maenaka, and Masafumi Takiguchi (*Equal contribution), Distinct HIV-1 Escape Patterns Selected by Cytotoxic T Cells with Identical Epitope Specificity, J. Virol. 87:2253-2263, 2013
Pol283-8-specific HLA-B*51:01-restricted CTLs play a critical role in long-term control of HIV-1. However, these CTLs select for the RT I135X escape mutation which may be accumulating in circulating HIV-1 sequences. We investigated the selection of the I135X mutation by CTLs specific for the same epitope but restricted by HLA-B*52:01. We found that Pol283-8-specific HLA-B*52:01-restricted CTLs were predominantly elicited in chronically HIV-1-infected individuals. These CTLs had a strong ability to suppress the replication of wild-type HIV-1, though this ability was weaker than that of HLA-B*51:01-restricted CTLs. The crystal structure of the HLA-B*52:01-Pol283-8 peptide complex provided clear evidence that HLA-B*52:01 presents the peptide similarly as HLA-B*51:01, ensuring the cross-presentation of this epitope by both alleles. Population-level analyses revealed a strong association of HLA-B*51:01 with the I135T mutant and a relatively weaker association of HLA-B*52:01 with several I135X mutants in both Japanese and predominantly Caucasian cohorts. An in vitro viral suppression assay revealed that the HLA-B*52:01-restricted CTLs failed to suppress the replication of the I135X mutant viruses, indicating the selection of these mutants by the CTLs. These results suggest that the different pattern of I135X mutant selection may have resulted from the difference between these 2 CTLs in their ability to suppress HIV-1 replication.