Distinct roles for signals relayed through the common cytokine receptor gamma chain and interleukin 7 receptor alpha chain in natural T cell development

Abstract

The commitment, differentiation, and expansion of mainstream alpha/beta T cells during ontogeny depend on the highly controlled interplay of signals relayed by cytokines through their receptors on progenitor cells. The role of cytokines in the development of natural killer (NK)1(+) natural T cells is less clearly understood. In an approach to define the role of cytokines in the commitment, differentiation, and expansion of NK1(+) T cells, their development was studied in common cytokine receptor gamma chain (gammac) and interleukin (IL)-7 receptor alpha (IL-7Ralpha)-deficient mice. These mutations block mainstream alpha/beta T cell ontogeny at an early prethymocyte stage. Natural T cells do not develop in gammac-deficient mice; they are absent in the thymus and peripheral lymphoid organs such as the liver and the spleen. In contrast, NK1(+) T cells develop in IL-7Ralpha-deficient mice in the thymus, and they are present in the liver and in the spleen. However, the absolute number of NK1(+) T cells in the thymus of IL-7Ralpha-deficient mice is reduced to approximately 10%, compared to natural T cell number in the wild-type thymus. Additional data revealed that NK1(+) T cell ontogeny is not impaired in IL-2- or IL-4-deficient mice, suggesting that neither IL-2, IL-4, nor IL-7 are required for their development. From these data, we conclude that commitment and/or differentiation to the NK1(+) natural T cell lineage requires signal transduction through the gammac, and once committed, their expansion requires signals relayed through the IL-7Ralpha.

Figure 1

Common cytokine receptor γc-deficient mice do not develop thymic NTα/β cells. Dot plots displaying CD44⁺ NKR-P1⁺ and CD44⁺TCR-α/β⁺ T cells among HSA^low CD8^low thymocytes of B6.γc^0/Y (seventh generation backcross to C57BL/6) and B6.IL-2Rγc⁺/^Y littermates. HSA^lowCD8^low thymocyte population was electronically gated and CD44^highNKR-P1⁺ T cells were analyzed with a FACScan^® flow cytometer.

Figure 2

IL-2–, IL-4–, and IL-7Rα–deficient mice develop thymic NTα/β T cells. (A) Dot plots of CD44^highNKR-P1⁺, CD44^highTCR-α/β^med, and CD44^highTCR-β8.1,8.2^med T cells among HSA^lowCD8^low thymocytes of B6.IL-2^0/0 (n = 5), B6.IL-4^0/0 (n = 6) and B6.IL-7Rα^0/0 mice (n = 6). (B) Dot plots of Ly6C^highNKR-P1⁺ T cells among HSA^lowCD8^low thymocytes. (C) Dot plots of IL-2 Rβ⁺NKR-P1⁺ (B6.IL-4^0/0 and B6.IL-7Rα^0/0) and of IL-2Rβ⁺TCR-α/β⁺ (B6.IL-2^0/0 and B6.IL-7Rα^0/0) T cells among HSA^lowCD8^low thymocytes. In B and C, the percentage of NT cells was almost equal in B6.IL-4^0/0, about half in B6.IL-7Rα^0/0, or up to twofold greater in B6.IL-2^0/0 compared with those in the wild type. (D) Absolute numbers of HSA^lowCD8^low thymocytes calculated as the thymocyte number times the fraction of this subset. (E) Thymic NT cells in wild-type, IL-2^0/0, IL-4^0/0 and IL-7Rα^0/0 mice. NT cell number was calculated from the percentages of the double-positive CD44⁺ and NKR-P1⁺, TCR-α/β⁺ or Vβ8.1,8.2⁺ thymocytes within the electronically gated HSA^lowCD8^low population in D as described previously (5).

Figure 2

IL-2–, IL-4–, and IL-7Rα–deficient mice develop thymic NTα/β T cells. (A) Dot plots of CD44^highNKR-P1⁺, CD44^highTCR-α/β^med, and CD44^highTCR-β8.1,8.2^med T cells among HSA^lowCD8^low thymocytes of B6.IL-2^0/0 (n = 5), B6.IL-4^0/0 (n = 6) and B6.IL-7Rα^0/0 mice (n = 6). (B) Dot plots of Ly6C^highNKR-P1⁺ T cells among HSA^lowCD8^low thymocytes. (C) Dot plots of IL-2 Rβ⁺NKR-P1⁺ (B6.IL-4^0/0 and B6.IL-7Rα^0/0) and of IL-2Rβ⁺TCR-α/β⁺ (B6.IL-2^0/0 and B6.IL-7Rα^0/0) T cells among HSA^lowCD8^low thymocytes. In B and C, the percentage of NT cells was almost equal in B6.IL-4^0/0, about half in B6.IL-7Rα^0/0, or up to twofold greater in B6.IL-2^0/0 compared with those in the wild type. (D) Absolute numbers of HSA^lowCD8^low thymocytes calculated as the thymocyte number times the fraction of this subset. (E) Thymic NT cells in wild-type, IL-2^0/0, IL-4^0/0 and IL-7Rα^0/0 mice. NT cell number was calculated from the percentages of the double-positive CD44⁺ and NKR-P1⁺, TCR-α/β⁺ or Vβ8.1,8.2⁺ thymocytes within the electronically gated HSA^lowCD8^low population in D as described previously (5).

Figure 2

IL-2–, IL-4–, and IL-7Rα–deficient mice develop thymic NTα/β T cells. (A) Dot plots of CD44^highNKR-P1⁺, CD44^highTCR-α/β^med, and CD44^highTCR-β8.1,8.2^med T cells among HSA^lowCD8^low thymocytes of B6.IL-2^0/0 (n = 5), B6.IL-4^0/0 (n = 6) and B6.IL-7Rα^0/0 mice (n = 6). (B) Dot plots of Ly6C^highNKR-P1⁺ T cells among HSA^lowCD8^low thymocytes. (C) Dot plots of IL-2 Rβ⁺NKR-P1⁺ (B6.IL-4^0/0 and B6.IL-7Rα^0/0) and of IL-2Rβ⁺TCR-α/β⁺ (B6.IL-2^0/0 and B6.IL-7Rα^0/0) T cells among HSA^lowCD8^low thymocytes. In B and C, the percentage of NT cells was almost equal in B6.IL-4^0/0, about half in B6.IL-7Rα^0/0, or up to twofold greater in B6.IL-2^0/0 compared with those in the wild type. (D) Absolute numbers of HSA^lowCD8^low thymocytes calculated as the thymocyte number times the fraction of this subset. (E) Thymic NT cells in wild-type, IL-2^0/0, IL-4^0/0 and IL-7Rα^0/0 mice. NT cell number was calculated from the percentages of the double-positive CD44⁺ and NKR-P1⁺, TCR-α/β⁺ or Vβ8.1,8.2⁺ thymocytes within the electronically gated HSA^lowCD8^low population in D as described previously (5).

Figure 3

NTα/β cells do not develop in peripheral lymphoid organs of γc-deficient mice, but develop in IL-7Rα^0/0 and IL-2^0/0 animals. Dot plots displaying NKR-P1⁺TCR-α/β⁺ cells among mononuclear cells isolated from the liver (A) and spleen (B) of wild-type and mutant mice. Liver NTα/β cells were stained with anti–NKR-P1-PE and anti–TCR-β-biotin (B6.γc^0/Y and B6.IL-7Rα^0/0) or anti–TCR-β-PE and anti–NKR-P1-biotin (B6.IL-2^+/0 and B6.IL-2^0/0). Splenic NTα/β cells were stained with anti–B220-FITC, anti-TCR-β-PE and anti-NKR-P1-biotin, and identified among B220^null splenocytes. In all cases, the biotinylated antibodies were detected by staining with streptavidin-RED670. The staining pattern of intrahepatic NT cells was observed in over 20 different preparations (see also reference 4).

Figure 3

NTα/β cells do not develop in peripheral lymphoid organs of γc-deficient mice, but develop in IL-7Rα^0/0 and IL-2^0/0 animals. Dot plots displaying NKR-P1⁺TCR-α/β⁺ cells among mononuclear cells isolated from the liver (A) and spleen (B) of wild-type and mutant mice. Liver NTα/β cells were stained with anti–NKR-P1-PE and anti–TCR-β-biotin (B6.γc^0/Y and B6.IL-7Rα^0/0) or anti–TCR-β-PE and anti–NKR-P1-biotin (B6.IL-2^+/0 and B6.IL-2^0/0). Splenic NTα/β cells were stained with anti–B220-FITC, anti-TCR-β-PE and anti-NKR-P1-biotin, and identified among B220^null splenocytes. In all cases, the biotinylated antibodies were detected by staining with streptavidin-RED670. The staining pattern of intrahepatic NT cells was observed in over 20 different preparations (see also reference 4).