Absence of Prr7 does not affect the susceptibility of mice to EAE.

<p>(A) Clinical EAE scores in 8–12 week old WT and Prr7 knockout female mice upon immunization with MOG peptide over time. (B) Cumulative EAE score from (A). (C) Maximum EAE scores from (A). Data are represented as mean +/- SEM of 13 (Prr7^-/-) and 14 (Prr7^+/+) mice per group from two independent experiments. n.s. not significant.</p

TCR response and AICD is unaffected in T cells from Prr7-deficient mice.

<p>(A) Flow cytometry analysis of the activation marker CD69 in Prr7-deficient CD4⁺ or CD8⁺ T cells stimulated with 1 μg of plate-bound anti-CD3 for 24 or 36 h. (B) Proliferation of Prr7^+/+ and Prr7^-/- splenocytes in response to TCR stimulation with plate-bound anti-CD3 measured as [³H]thymidine incorporation (DNA synthesis). cpm, counts per minute. (C) Schema of the <i>in vitro</i> AICD induction protocol. (D) Representative examples of flow cytometry analysis of AICD in T cells upon restimulation. Live = PI^-Annexin V^-, Apoptotic = PI^-Annexin V⁺, Dead = PI⁺Annexin V⁺ (E) Quantification and statistical analysis of AICD performed as shown in (D). (F) Immunoblotting of c-Jun total levels in restimulated T cells isolated from three different wild-type and knockout mice (#1, #2, #3). Tubulin served as a loading control. Data in (A, B, D) represent the mean + SEM of at least three animals per group. n.s., not significant.</p

Prr7 deficiency does not influence T cell response to <i>Listeria monocytogenes</i> infection.

<p>(A) <i>Prr7</i>^-/- mice and <i>Prr7</i>^+/+ control mice were i.v. infected with 1x10⁴ Lm ova. On day 9 post infection, colony forming units were determined in spleen and liver. (B) Representative dot plot of CD4⁺ and CD8⁺ T cells in the spleen of infected mice. (C) Frequency of CD4⁺ and CD8⁺ T cells in the spleen of infected mice. (D) Absolute number of CD4⁺ and CD8⁺ T cells in the spleen of infected mice. (E) Frequency of CD8⁺ naive (CD62L⁺CD44^-), activated (CD62L^-CD44⁺) and memory (CD62L⁺CD44⁺) T cells in the spleen of infected mice. (F) Absolute number of CD8⁺ naive, activated and memory T cells in the spleen of infected mice. (G-K) Splenocytes of infected mice were restimulated with Ova_257-264-peptide (SIINFEKL, 10⁻⁸ M) or left unstimulated for 12 h in the presence of Brefeldin A, to allow for the intracellular accumulation of cytokines. (G) Dot plot of IFN-γ and TNF-producing CD8⁺ T cells without restimulation. (H) Dot plot of TNF-producing CD8⁺ T cells after restimulation. (I) Dot plot of IFN-γ producing CD8⁺ T cells after restimulation. (J) Frequency of IFN-γ and TNF-producing CD8⁺ T cells in the spleen of infected mice. (K) Absolute number of IFN-γ and TNF-producing CD8⁺ T cells in the spleen of infected mice. Data are represented as mean ± SEM of 4–5 mice per group. n.s. not significant.</p

T cell development is largely unaffected in Prr7-deficient mice.

<p>(A) Total numbers of nucleated cells in the thymus (left) and spleen (right) isolated from Prr7^+/+ and Prr7^-/- mice as counted using a haemocytometer. (B) Schematic representation of T cell developmental stages in the thymus. DN, double negative, DP, double positive, SP, single positive. Lower panels with dot plots are representative examples of flow cytometry analysis of thymocyte subpopulations. Percentages of DN subpopulations (C), DP subpopulations (D), and SP subpopulations (E) in thymi of Prr7^+/+ and Prr7^-/- mice as analysed by flow cytometry. (F) Flow cytometry analysis of CD4⁺ and CD8⁺ T cells subpopulations in the secondary lymphatic organs spleen and lymph nodes expressed as percentage of total. (G) Flow cytometry analysis of splenic CD3⁺ T cells expressing markers of naïve T cells (CD62⁺CD25^-), activated T cells (CD62L^-CD25⁺), or memory T cells (CD62L^-CD25⁺). Data in (A-G) represent the mean +SEM, n = 3–8. *p < 0.05, n.s., not significant.</p

Table_1_Self-reactivity of CD8 T-cell clones determines their differentiation status rather than their responsiveness in infections.xlsx

Mature T cells are selected for recognizing self-antigens with low to intermediate affinity in the thymus. Recently, the relative differences in self-reactivity among individual T-cell clones were appreciated as important factors regulating their fate and immune response, but the role of self-reactivity in T-cell biology is incompletely understood. We addressed the role of self-reactivity in T-cell diversity by generating an atlas of mouse peripheral CD8+ T cells, which revealed two unconventional populations of antigen-inexperienced T cells. In the next step, we examined the steady-state phenotype of monoclonal T cells with various levels of self-reactivity. Highly self-reactive clones preferentially differentiate into antigen-inexperienced memory-like cells, but do not form a population expressing type I interferon-induced genes, showing that these two subsets have unrelated origins. The functional comparison of naïve monoclonal CD8+ T cells specific to the identical model antigen did not show any correlation between the level of self-reactivity and the magnitude of the immune response.</p

DataSheet_1_Self-reactivity of CD8 T-cell clones determines their differentiation status rather than their responsiveness in infections.pdf

Mature T cells are selected for recognizing self-antigens with low to intermediate affinity in the thymus. Recently, the relative differences in self-reactivity among individual T-cell clones were appreciated as important factors regulating their fate and immune response, but the role of self-reactivity in T-cell biology is incompletely understood. We addressed the role of self-reactivity in T-cell diversity by generating an atlas of mouse peripheral CD8+ T cells, which revealed two unconventional populations of antigen-inexperienced T cells. In the next step, we examined the steady-state phenotype of monoclonal T cells with various levels of self-reactivity. Highly self-reactive clones preferentially differentiate into antigen-inexperienced memory-like cells, but do not form a population expressing type I interferon-induced genes, showing that these two subsets have unrelated origins. The functional comparison of naïve monoclonal CD8+ T cells specific to the identical model antigen did not show any correlation between the level of self-reactivity and the magnitude of the immune response.</p