Tat does not contribute to the control of HSV1 acute infection.

<p>Control and Tat-treated HSV1-infected C57/BL6 mice were checked daily for the appearance of disease signs. (<b>A</b>) Mean of disease scores ± SEM of 20 mice per group is shown. For statistical analysis two-tailed Mann Whitney test was used. **P<0.01. (<b>B</b>) Probability of developing disease signs is shown for each group. Figure represents Kaplan-Meier estimation of the probability of clinical manifestations. For statistical analysis Log rank test was used. One representative experiment out of three is shown.</p

Tat-mediated CD8⁺ T cell activation.

<p>(<b>A</b>) Fresh splenocytes from C57BL/6 mice were co-cultured, in the presence or absence of 1µg/ml of Tat, with autologous splenocytes previously loaded with the SSI peptide epitope. After 8 days, cells were assayed in IFNγ Elispot. One representative experiment out of five is shown. (<b>B</b>) Splenocytes were purified from HSV1-infected C57BL/6 mice at day 8 post-infection and assayed in IFNγ Elispot against the SSI epitope in the presence or absence of 1 µg/ml of Tat. One representative experiment out of five is shown. (<b>C</b>) Balb/c mice (3 per groups) were injected with 5 µg of Gag alone or in combination with 5 µg of Tat. Ten days after vaccination mice were sacrificed and fresh splenocytes assayed in IFNγ Elispot against the indicated Gag-derived T cell peptide epitopes (see also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0077746#pone.0077746.s002" target="_blank">Table S1</a>). One representative experiment out of three is shown.</p

Tat administered at the time of antigen-priming favors an effector memory phenotype.

<p>Control and Tat-treated mice were infected with HSV1 wt (<b>A</b> and <b>B</b>) or with replicative-defective HSV1 (<b>C</b> and <b>D</b>) and sacrificed at days 70 post-infection. (<b>A</b> and <b>C</b>) Percentage of SSI-specific CD8⁺ T cells detected by dextramer staining. (<b>B</b> and <b>D</b>) CD62L expression was measured by flow cytometry on SSI-specific CD8⁺ T cells. Data are presented as mean ± SEM of 5 mice per group. For statistical analysis two-tailed Mann Whitney test was used. *P<0.05. One representative experiment out of three is shown.</p

Tat does not activate bystander T cells.

<p>Control and Tat-treated HSV1-infected C57/BL6 mice were sacrificed at days 4, 6, 8, 10 and 13 post-infection. CD8⁺ (<b>A</b>), CD4⁺ (<b>C</b>) and B (<b>D</b>) lymphocytes numbers were measured by flow cytometry. Data are presented as mean ± SEM of 5 mice per group. For statistical analysis two-tailed Mann Whitney test was used. *P<0.05. One representative experiment out of three is shown. (<b>B</b>) CD95 expression was measured by flow cytometry on CD8⁺ T cells. One representative experiment out of five is shown.</p

Tat modulates the kinetics and the magnitude of CTL responses.

<p>Splenocytes were purified from control and Tat-treated HSV1-infected C57/BL6 mice at days 4, 5, 6, 7, 8, 10 and 13 post-infection. (<b>A</b>) Percentage of SSI-specific CD8⁺ T cells detected by dextramer staining. Data are presented as mean ± SEM of 5 mice per group. One representative experiment out of three is shown. (<b>B</b>) Kinetics of SSI- and QTF-specific cellular responses detected by IFNγ Elispot on fresh splenocytes. Data are presented as mean ± SEM of 5 mice per group (left panel). Expansion and contraction were normalized, for each group, to values detected at day 7 (right panel). One representative experiment out of three is shown. (<b>C</b>) SSI- and QTF- specific IFNγ responses at days 5 and 6 post-infection. (<b>D</b>) SSI- and QTF- specific IFNγ responses at day 7 post-infection. (<b>E</b>) SSI- and QTF-specific IFNγ responses at day 13 post-infection. For statistical analysis two-tailed Mann Whitney test was used. *P<0.05. </p

Tat administered at the time of antigen-priming favors a Th1 profile of the humoral response.

<p>Blood samples from control and Tat-treated HSV1-infected mice were collected and the presence of anti-HSV1 antibodies was detected by ELISA assay. (<b>A</b>) Anti-HSV1 IgG1 and IgG2a were measured at days 20 (left) and 70 (right) post-infection. (<b>B</b>) Total anti-HSV1 IgG were measured at days 20 (left) and 70 (right) post-infection. Data are presented as mean ± SEM of 5 mice per group. For statistical analysis two-tailed Mann Whitney test was used. *P<0.05. One representative experiment out of three is shown.</p