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GITR intrinsically sustains early type 1 and late follicular helper CD4 T cell accumulation to control a chronic viral infection.

Clouthier DL, Zhou AC, Wortzman ME, Luft O, Levy GA, Watts TH - PLoS Pathog. (2015)

Bottom Line: While several studies have addressed the role of inhibitory receptors and soluble factors such as PD-1 and IL-10, significantly less work has addressed the role of T cell co-stimulatory molecules during chronic viral infection.GITR-dependent phosphorylation of the p65 subunit of NF-κB as well as phosphorylation of the downstream mTORC1 target, S6 ribosomal protein, were detected at day three post-infection (p.i.), and defects in CD4 T cell accumulation in GITR-deficient T cells were apparent starting at day five p.i.GITR also increases the ratio of T follicular helper to T follicular regulatory cells and thereby enhances LCMV-specific IgG production.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, University of Toronto, Toronto, Ontario, Canada.

ABSTRACT
CD4 T cells are critical for control of persistent infections; however, the key signals that regulate CD4 T help during chronic infection remain incompletely defined. While several studies have addressed the role of inhibitory receptors and soluble factors such as PD-1 and IL-10, significantly less work has addressed the role of T cell co-stimulatory molecules during chronic viral infection. Here we show that during a persistent infection with lymphocytic choriomeningitis virus (LCMV) clone 13, mice lacking the glucocorticoid-induced tumor necrosis factor receptor related protein (GITR) exhibit defective CD8 T cell accumulation, increased T cell exhaustion and impaired viral control. Differences in CD8 T cells and viral control between GITR+/+ and GITR-/- mice were lost when CD4 T cells were depleted. Moreover, mixed bone marrow chimeric mice, as well as transfer of LCMV epitope-specific CD4 or CD8 T cells, demonstrated that these effects of GITR are largely CD4 T cell-intrinsic. GITR is dispensable for initial CD4 T cell proliferation and differentiation, but supports the post-priming accumulation of IFNγ+IL-2+ Th1 cells, facilitating CD8 T cell expansion and early viral control. GITR-dependent phosphorylation of the p65 subunit of NF-κB as well as phosphorylation of the downstream mTORC1 target, S6 ribosomal protein, were detected at day three post-infection (p.i.), and defects in CD4 T cell accumulation in GITR-deficient T cells were apparent starting at day five p.i. Consistently, we pinpoint IL-2-dependent CD4 T cell help for CD8 T cells to between days four and eight p.i. GITR also increases the ratio of T follicular helper to T follicular regulatory cells and thereby enhances LCMV-specific IgG production. Together, these findings identify a CD4 T cell-intrinsic role for GITR in sustaining early CD8 and late humoral responses to collectively promote control of chronic LCMV clone 13 infection.

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The effect of GITR on the CD8 T cell response is CD8 T cell-extrinsic.(A) Schematic indicating that 103 GITR+/+ or GITR-/- CD45.1 P14 cells from F2 littermates were adoptively transferred into separate naïve CD45.2 congenic mice one day prior to infection with 2×106 ffu LCMV cl 13. (B and C) At day eight p.i., the frequency of CD45.1 P14 cells in peripheral blood (B, left) and the number of total (B, right) and IFNγ+CD107a+TNF+ (C) P14 cells were determined. (D, E) CD45.1 P14 cells were evaluated for expression of PD-1 and Tim-3, with summary plots and representative stains shown. (F) Viral load in spleen and kidney was evaluated from mice at day eight p.i. Broken horizontal lines represent the limit of detection. Each symbol represents an individual mouse, with bars indicating mean ± SEM. Data are representative of four experiments with at least four mice per group, with two repeats in the blood and two repeats in the spleen and kidney.
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ppat.1004517.g003: The effect of GITR on the CD8 T cell response is CD8 T cell-extrinsic.(A) Schematic indicating that 103 GITR+/+ or GITR-/- CD45.1 P14 cells from F2 littermates were adoptively transferred into separate naïve CD45.2 congenic mice one day prior to infection with 2×106 ffu LCMV cl 13. (B and C) At day eight p.i., the frequency of CD45.1 P14 cells in peripheral blood (B, left) and the number of total (B, right) and IFNγ+CD107a+TNF+ (C) P14 cells were determined. (D, E) CD45.1 P14 cells were evaluated for expression of PD-1 and Tim-3, with summary plots and representative stains shown. (F) Viral load in spleen and kidney was evaluated from mice at day eight p.i. Broken horizontal lines represent the limit of detection. Each symbol represents an individual mouse, with bars indicating mean ± SEM. Data are representative of four experiments with at least four mice per group, with two repeats in the blood and two repeats in the spleen and kidney.

Mentions: Previous work has shown an intrinsic role for GITR in sustaining the survival of TCR transgenic CD8 T cells during acute influenza virus infection [26]. To determine if CD8 T cell-intrinsic effects were also responsible for defects in LCMV cl 13 control in GITR-/- mice, we crossed GITR-/- with P14 mice, which have a transgenic TCR specific for the Db-restricted LCMV GP33–41 epitope [27]. We transferred 103 purified CD8 T cells from CD45.1 GITR+/+ or GITR-/- P14 littermates into CD45.2 congenic mice one day prior to LCMV cl 13 infection (Fig. 3A). P14 T cells lacking GITR showed no impairment in expansion, and in fact showed a slight increase in frequency (Fig. 3B). There was also no difference in IFNγ production or degranulation, PD-1 or Tim-3 expression between the GITR-sufficient or GITR-deficient P14 T cells (Fig. 3, C–E). Moreover, the absence of GITR only on the transferred P14 T cells had no effect on viral control at day eight p.i. (Fig. 3F). Thus, the effect of GITR in potentiating the CD8 T cell response to LCMV cl 13 is CD8 T cell-extrinsic.


GITR intrinsically sustains early type 1 and late follicular helper CD4 T cell accumulation to control a chronic viral infection.

Clouthier DL, Zhou AC, Wortzman ME, Luft O, Levy GA, Watts TH - PLoS Pathog. (2015)

The effect of GITR on the CD8 T cell response is CD8 T cell-extrinsic.(A) Schematic indicating that 103 GITR+/+ or GITR-/- CD45.1 P14 cells from F2 littermates were adoptively transferred into separate naïve CD45.2 congenic mice one day prior to infection with 2×106 ffu LCMV cl 13. (B and C) At day eight p.i., the frequency of CD45.1 P14 cells in peripheral blood (B, left) and the number of total (B, right) and IFNγ+CD107a+TNF+ (C) P14 cells were determined. (D, E) CD45.1 P14 cells were evaluated for expression of PD-1 and Tim-3, with summary plots and representative stains shown. (F) Viral load in spleen and kidney was evaluated from mice at day eight p.i. Broken horizontal lines represent the limit of detection. Each symbol represents an individual mouse, with bars indicating mean ± SEM. Data are representative of four experiments with at least four mice per group, with two repeats in the blood and two repeats in the spleen and kidney.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4295864&req=5

ppat.1004517.g003: The effect of GITR on the CD8 T cell response is CD8 T cell-extrinsic.(A) Schematic indicating that 103 GITR+/+ or GITR-/- CD45.1 P14 cells from F2 littermates were adoptively transferred into separate naïve CD45.2 congenic mice one day prior to infection with 2×106 ffu LCMV cl 13. (B and C) At day eight p.i., the frequency of CD45.1 P14 cells in peripheral blood (B, left) and the number of total (B, right) and IFNγ+CD107a+TNF+ (C) P14 cells were determined. (D, E) CD45.1 P14 cells were evaluated for expression of PD-1 and Tim-3, with summary plots and representative stains shown. (F) Viral load in spleen and kidney was evaluated from mice at day eight p.i. Broken horizontal lines represent the limit of detection. Each symbol represents an individual mouse, with bars indicating mean ± SEM. Data are representative of four experiments with at least four mice per group, with two repeats in the blood and two repeats in the spleen and kidney.
Mentions: Previous work has shown an intrinsic role for GITR in sustaining the survival of TCR transgenic CD8 T cells during acute influenza virus infection [26]. To determine if CD8 T cell-intrinsic effects were also responsible for defects in LCMV cl 13 control in GITR-/- mice, we crossed GITR-/- with P14 mice, which have a transgenic TCR specific for the Db-restricted LCMV GP33–41 epitope [27]. We transferred 103 purified CD8 T cells from CD45.1 GITR+/+ or GITR-/- P14 littermates into CD45.2 congenic mice one day prior to LCMV cl 13 infection (Fig. 3A). P14 T cells lacking GITR showed no impairment in expansion, and in fact showed a slight increase in frequency (Fig. 3B). There was also no difference in IFNγ production or degranulation, PD-1 or Tim-3 expression between the GITR-sufficient or GITR-deficient P14 T cells (Fig. 3, C–E). Moreover, the absence of GITR only on the transferred P14 T cells had no effect on viral control at day eight p.i. (Fig. 3F). Thus, the effect of GITR in potentiating the CD8 T cell response to LCMV cl 13 is CD8 T cell-extrinsic.

Bottom Line: While several studies have addressed the role of inhibitory receptors and soluble factors such as PD-1 and IL-10, significantly less work has addressed the role of T cell co-stimulatory molecules during chronic viral infection.GITR-dependent phosphorylation of the p65 subunit of NF-κB as well as phosphorylation of the downstream mTORC1 target, S6 ribosomal protein, were detected at day three post-infection (p.i.), and defects in CD4 T cell accumulation in GITR-deficient T cells were apparent starting at day five p.i.GITR also increases the ratio of T follicular helper to T follicular regulatory cells and thereby enhances LCMV-specific IgG production.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, University of Toronto, Toronto, Ontario, Canada.

ABSTRACT
CD4 T cells are critical for control of persistent infections; however, the key signals that regulate CD4 T help during chronic infection remain incompletely defined. While several studies have addressed the role of inhibitory receptors and soluble factors such as PD-1 and IL-10, significantly less work has addressed the role of T cell co-stimulatory molecules during chronic viral infection. Here we show that during a persistent infection with lymphocytic choriomeningitis virus (LCMV) clone 13, mice lacking the glucocorticoid-induced tumor necrosis factor receptor related protein (GITR) exhibit defective CD8 T cell accumulation, increased T cell exhaustion and impaired viral control. Differences in CD8 T cells and viral control between GITR+/+ and GITR-/- mice were lost when CD4 T cells were depleted. Moreover, mixed bone marrow chimeric mice, as well as transfer of LCMV epitope-specific CD4 or CD8 T cells, demonstrated that these effects of GITR are largely CD4 T cell-intrinsic. GITR is dispensable for initial CD4 T cell proliferation and differentiation, but supports the post-priming accumulation of IFNγ+IL-2+ Th1 cells, facilitating CD8 T cell expansion and early viral control. GITR-dependent phosphorylation of the p65 subunit of NF-κB as well as phosphorylation of the downstream mTORC1 target, S6 ribosomal protein, were detected at day three post-infection (p.i.), and defects in CD4 T cell accumulation in GITR-deficient T cells were apparent starting at day five p.i. Consistently, we pinpoint IL-2-dependent CD4 T cell help for CD8 T cells to between days four and eight p.i. GITR also increases the ratio of T follicular helper to T follicular regulatory cells and thereby enhances LCMV-specific IgG production. Together, these findings identify a CD4 T cell-intrinsic role for GITR in sustaining early CD8 and late humoral responses to collectively promote control of chronic LCMV clone 13 infection.

Show MeSH
Related in: MedlinePlus