Summary of the publications
2007-2008
Takamasa Ueno, Chihiro Motozono, Sachi Douki, Philip Mwimanzi, Susanne Rauch, Oliver T. Fackler, Shinichi Oka, and Masafumi Takiguchi, Cytotoxic T lymphocyte-mediated selective pressure influences dynamic evolution and pathogenic functions of HIV-1 Nef , J. Immunol. 180:1107-1116, 2008
HIV-1 Nef plays multiple roles in modulating immune responses, even though it is a dominant CTL target itself. How Nef accomplishes the balance between such conflicting selective pressures remains elusive. By genetic and functional studies, we found that Arg75Thr and Tyr85Phe mutations, located in a well-conserved proline-rich region in Nef, were differently associated with escape from CTL responses specific for two overlapping HLA-B35-restricted epitopes. CTLs specific for an epitope, that selected Tyr85Phe, were elicited earlier and had more potent functional avidities than did those that selected Arg75Thr. Although the double mutant could escape from both CTLs, the mutations are rarely observed in combination naturally. Introduction of both mutations reduced Nef’s HLA class I down-regulation activity and increased the susceptibility of virus-infected cells to recognition by CTLs targeting other epitopes. Moreover, the mutant Nef was impaired in the association with activated cellular kinases and in the enhancement of viral replication. These results highlight CTL immunosurveillance as important modulators of Nef’s biological activity in the infected host.
Mamoru Fujiwara, Junko Tanuma, Hirokazu Koizumi, Yuka Kawashima, Kazutaka Honda, Saori Mastuoka-Aizawa, Sachi Dohki, Shinichi Oka and Masafumi Takiguchi, Different Ability of Escape Mutant-Specific Cytotoxic T Cells to Suppress Replication of Escape Mutant and Wild-type HIV-1 in New Hosts, J. Virol., 82: 138-147, 2008
There is much evidence that in human immunodeficiency virus type 1 (HIV-1)-infected individuals, strong cytotoxic T lymphocyte (CTL)-mediated immune pressure results in the selection of HIV-1 mutants that have escaped from wild-type-specific CTLs. If escape mutant-specific CTLs are not elicited in new hosts sharing donor HLA molecules, the transmission of these mutants results in the accumulation of escape mutants in the population. However, whether escape mutant-specific CTLs are definitively not elicited in new hosts sharing donor HLA molecules still remains unclear. A previous study showed that a Y-to-F substitution at the second position (2F) of the Nef138-10 epitope is significantly detected in HLA-A*2402+ hemophilic donors. Presently, we confirmed that this 2F mutant was an escape mutant by demonstrating strong and weak abilities of Nef138-10-specific CTL clones to suppress replication of the wild-type and 2F mutant viruses, respectively. We demonstrated the existence of the 2F-specific CTLs in three new hosts who had been primarily infected with the 2F mutant. The 2F-specific CTL clones suppressed the replication of both wild-type and mutant viruses. However, the abilities of these clones to suppress replication of the 2F virus were much weaker than those of wild-type-specific and the 2F-specific ones to suppress replication of the wild-type virus. These findings indicate that the 2F mutant is conserved in HIV-1-infected donors having HLA-A*2402, because the 2F-specific CTLs failed to completely suppress the 2F mutant replication and effectively prevented viral reversion in new hosts carrying HLA-A*2402.
Center for AIDS Research Best Paper Award 2007
Takamasa Ueno, Yuka Idegami, Chihiro Motozono, Shinichi Oka and Masafumi Takiguchi, Altering effects of antigenic variations in HIV-1 on antiviral effectiveness of HIV-specific CTLs J. Immunol. 178: 5513-5523, 2007
The mutational escape of HIV-1 from established CTL responses is becoming evident. However, it is not yet clear whether antigenic variations of HIV-1 may have an additional effect on the differential antiviral effectiveness of HIV-specific CTLs. Herein, we characterized HIV-specific CTL responses toward Pol, Env, and Nef optimal epitopes presented by HLA-B*35 during a chronic phase of HIV-1 infection. We found CTL escape variants within Pol and Nef epitopes that affected recognition by TCRs, although there was no mutation within the Env epitope. An analysis of peptide-HLA tetrameric complexes revealed that CD8 T cells exclusively specific for the Nef variant were generated following domination by the variant viruses. The variant-specific cells were capable of killing target cells and producing antiviral cytokines but showed impaired Ag-specific proliferation ex vivo, whereas wild-type specific cells had potent activities. Moreover, clonotypic CD8 T cells specific for the Pol variant showed diminished proliferation, whereas Env-specific ones had no functional heterogeneity. Taken together, our data indicate that antigenic variations that abolished TCR recognition not only resulted in escape from established CTL responses but also eventually generated another subset of variant-specific CTLs having decreased antiviral activity, causing an additional negative effect on antiviral immune responses during a chronic HIV infection.
Mamoru Fujiwara and Masafumi Takiguchi, HIV-1-Specific CTLs Effectively Suppress Replication of HIV-1 in HIV-1-infected Macrophages, Blood 109:4832-4838, 2007
Both CD4+ T cells and macrophages are major reservoirs of HIV-1. Previous study showed that HIV-1-specific cytolytic T lymphocytes (CTLs) hardly recognize HIV- 1-infected CD4+ T cells because of Nefmediated HLA class I down-regulation, suggesting that HIV-1 escapes from HIV- 1-specific CTLs and continues to replicate in HIV-1-infected donors. On the other hand, the CTL recognition of HIV-1- infected macrophages and the effect of Nef-mediated HLA class I down-regulation on this recognition still remain unclear. We show a strong HIV-1 antigen presentation by HIV-1-infected macrophages. HIV-1-specific CTLs had strong abilities to suppress HIV-1R5 virus replication in HIV-1-infected macrophages and to kill HIV-1R5-infected macrophages. Nef-mediated HLA class I down-regulation minimally influenced the recognition of HIV-1-infected macrophages by HIV-1-specific CTLs. In addition, HIV-1-infected macrophages had a stronger ability to stimulate the proliferation of HIV-1-specific CTLs than HIV-1-infected CD4+ T cells. Thus, the effect of Nef-mediated HLA class I down-regulation was less critical with respect to the recognition by HIV-1-specific CTLs of HIV-infected macrophages than that of HIV-1-infected CD4+ T cells. These findings support the idea that the strong HIV-1 antigen presentation by HIV-1-infected macrophages is one of the mechanisms mediating effective induction of HIV-1-specific CTLs in the acute and early chronic phases of HIV-1 infection.
Takaaki Kondo, Hiroshi Takata and Masafumi Takiguchi, Functional Expression of Chemokine Receptor CCR6 on Human Effector Memory CD8+ T Cells, Eur. J. Immunol 37:54-65, 2007　
Since CCR6 is a receptor for the chemokine CCL20, which is produced in tissues such as intestine and colon, it is thought that T cells expressing CCR6 are involved in mucosal immunity. The expression and function of CCR6 on human CD8+ T cells have not well been analyzed, although it is known that this receptor is expressed on a subset of human CD8+ T cells. We here characterize human CCR6+ CD8+ T cells. Multi-color flow cytometric analysis demonstrated that CCR6+ cells are predominantly found among CD8+ T cells having the memory phenotype. The expression of CCR6 is positively and negatively correlated with that of CCR5 and CCR7, respectively. CCR6+ CD8+ T cells express granzyme A and a low level of perforin but not granzyme B. In addition, amajor population among these cells has the ability to produce IFN-c and TNF-a but not IL-2. These results indicate that CCR6+ CD8+ T cells have characteristics of early effector memory cells rather than effector or central memory cells. A chemotaxis assay revealed that CCR6+ CD8+ T cells have the ability to migrate in response to CCL20, suggesting that these Tcells migrate to tissues such as colon and are involved in mucosal immunity.

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