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Akt1-mediated Gata3 phosphorylation controls the repression of IFNγ in memory-type Th2 cells.

Hosokawa H, Tanaka T, Endo Y, Kato M, Shinoda K, Suzuki A, Motohashi S, Matsumoto M, Nakayama KI, Nakayama T - Nat Commun (2016)

Bottom Line: We also identify Akt1 as a Gata3-phosphorylating kinase, and the activation of Akt1 induces derepression of Tbx21 and Ifng expression in Th2 cells.Moreover, T-bet-dependent IFNγ expression in IFNγ-producing memory Th2 cells appears to be controlled by the phosphorylation status of Gata3 in human and murine systems.Thus, this study highlights the molecular basis for posttranslational modifications of Gata3 that control the regulation of IFNγ expression in memory Th2 cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670, Japan.

ABSTRACT
Th2 cells produce Th2 cytokines such as IL-4, IL-5 and IL-13, but repress Th1 cytokine IFNγ. Recent studies have revealed various distinct memory-type Th2 cell subsets, one of which produces a substantial amount of IFNγ in addition to Th2 cytokines, however it remains unclear precisely how these Th2 cells produce IFNγ. We herein show that phosphorylation of Gata3 at Ser308, Thr315 and Ser316 induces dissociation of a histone deacetylase Hdac2 from the Gata3/Chd4 repressive complex in Th2 cells. We also identify Akt1 as a Gata3-phosphorylating kinase, and the activation of Akt1 induces derepression of Tbx21 and Ifng expression in Th2 cells. Moreover, T-bet-dependent IFNγ expression in IFNγ-producing memory Th2 cells appears to be controlled by the phosphorylation status of Gata3 in human and murine systems. Thus, this study highlights the molecular basis for posttranslational modifications of Gata3 that control the regulation of IFNγ expression in memory Th2 cells.

No MeSH data available.


Related in: MedlinePlus

Increased phosphorylation status of Gata3 and Akt1 in IFNγ-producing memory Th2 cells.(a) Memory Th2 cells were stimulated with PMA plus ionomycin for 6 h. The cells were divided into four distinct subpopulations according to their expression of IL-4 and IFNγ (left). Intracellular staining profiles of T-bet in these four subpopulations are shown (middle). The number in the histogram represents the MFI. The grey-filled histogram shows isotype control staining. A summary of the MFI of T-bet in IFNγ-producing and -nonproducing cells is presented (right) (n=5). (b) Intracellular staining profiles of IFNγ and IL-4 in effector and memory Th2 cells from WT or Tbx21-deficient mice are shown with the percentages of cells in each area (left). A summary of the percentage of IFNγ-producing memory Th2 cells is presented (right) (n=3). (c) Intracellular staining profiles of Gata3, phospho-Gata3, Akt1 and phospho-Akt1 in IFNγ-producing and -nonproducing memory Th2 cells are shown (left). A summary of the MFI value of Gata3 and phospho-Gata3 and the ratio of the MFI (phospho-Gata3/total Gata3) in IFNγ-producing and IFNγ-nonproducing memory Th2 cells is presented (right) (n=5). (d) Intracellular staining profiles of IFNγ, Gata3, phospho-Gata3, T-bet and phospo-Akt1 are shown. The number represents the MFI of the y axis in each area (top). A summary of the MFI in each area (IFNγ: Low, Int, High) is presented (bottom) (n=5). (e) Memory Th2 cells were cultured in IL-7-containing medium with or without Akt inhibitor XI (10 μM) for 2 days. Intracellular staining profiles of IFNγ and IL-4 are shown (left). A summary of the percentage of IL-4- and IFNγ-producing memory Th2 cells is presented (right) (n=4). **P<0.01, *P<0.05 by Student's t-test. Five (a), four (e) and three (b,c,d) independent experiments were performed with similar results.
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f6: Increased phosphorylation status of Gata3 and Akt1 in IFNγ-producing memory Th2 cells.(a) Memory Th2 cells were stimulated with PMA plus ionomycin for 6 h. The cells were divided into four distinct subpopulations according to their expression of IL-4 and IFNγ (left). Intracellular staining profiles of T-bet in these four subpopulations are shown (middle). The number in the histogram represents the MFI. The grey-filled histogram shows isotype control staining. A summary of the MFI of T-bet in IFNγ-producing and -nonproducing cells is presented (right) (n=5). (b) Intracellular staining profiles of IFNγ and IL-4 in effector and memory Th2 cells from WT or Tbx21-deficient mice are shown with the percentages of cells in each area (left). A summary of the percentage of IFNγ-producing memory Th2 cells is presented (right) (n=3). (c) Intracellular staining profiles of Gata3, phospho-Gata3, Akt1 and phospho-Akt1 in IFNγ-producing and -nonproducing memory Th2 cells are shown (left). A summary of the MFI value of Gata3 and phospho-Gata3 and the ratio of the MFI (phospho-Gata3/total Gata3) in IFNγ-producing and IFNγ-nonproducing memory Th2 cells is presented (right) (n=5). (d) Intracellular staining profiles of IFNγ, Gata3, phospho-Gata3, T-bet and phospo-Akt1 are shown. The number represents the MFI of the y axis in each area (top). A summary of the MFI in each area (IFNγ: Low, Int, High) is presented (bottom) (n=5). (e) Memory Th2 cells were cultured in IL-7-containing medium with or without Akt inhibitor XI (10 μM) for 2 days. Intracellular staining profiles of IFNγ and IL-4 are shown (left). A summary of the percentage of IL-4- and IFNγ-producing memory Th2 cells is presented (right) (n=4). **P<0.01, *P<0.05 by Student's t-test. Five (a), four (e) and three (b,c,d) independent experiments were performed with similar results.

Mentions: Recently, distinct memory-type Th2 cell subsets have been identified in several groups, which produce a substantial amount of IL-5, IL-17 or IFNγ in addition to IL-4 and IL-13 (ref. 7). IFNγ expression in Th2 cells is reported to be dependent on T-bet and plays an important role in the protective immunity against viral infections6. We examined whether Gata3 phosphorylation-induced derepression of Tbx21 expression is involved in the IFNγ production from these memory-type Th2 cells. Antigen-specific memory Th2 cells can be efficiently generated in vivo by adoptive transfer of effector Th2 cells34. To determine the expression levels of T-bet in IFNγ-producing memory Th2 cells, the cells were divided into four distinct subpopulations according to their expression of IL-4 and IFNγ. Consistent with previous report35, we found that IFNγ-producing memory Th2 cells express higher levels of T-bet protein as compared with non-IFNγ-producing memory Th2 cells (Fig. 6a). To examine whether IFNγ production from memory Th2 cells is T-bet-dependent, memory Th2 cells from Tbx21-deficient cells were generated. While there was no difference in effector Th2 cell differentiation between WT and Tbx21-deficient cells, IFNγ production from Tbx21-deficient memory Th2 cells was strongly inhibited (Fig. 6b).


Akt1-mediated Gata3 phosphorylation controls the repression of IFNγ in memory-type Th2 cells.

Hosokawa H, Tanaka T, Endo Y, Kato M, Shinoda K, Suzuki A, Motohashi S, Matsumoto M, Nakayama KI, Nakayama T - Nat Commun (2016)

Increased phosphorylation status of Gata3 and Akt1 in IFNγ-producing memory Th2 cells.(a) Memory Th2 cells were stimulated with PMA plus ionomycin for 6 h. The cells were divided into four distinct subpopulations according to their expression of IL-4 and IFNγ (left). Intracellular staining profiles of T-bet in these four subpopulations are shown (middle). The number in the histogram represents the MFI. The grey-filled histogram shows isotype control staining. A summary of the MFI of T-bet in IFNγ-producing and -nonproducing cells is presented (right) (n=5). (b) Intracellular staining profiles of IFNγ and IL-4 in effector and memory Th2 cells from WT or Tbx21-deficient mice are shown with the percentages of cells in each area (left). A summary of the percentage of IFNγ-producing memory Th2 cells is presented (right) (n=3). (c) Intracellular staining profiles of Gata3, phospho-Gata3, Akt1 and phospho-Akt1 in IFNγ-producing and -nonproducing memory Th2 cells are shown (left). A summary of the MFI value of Gata3 and phospho-Gata3 and the ratio of the MFI (phospho-Gata3/total Gata3) in IFNγ-producing and IFNγ-nonproducing memory Th2 cells is presented (right) (n=5). (d) Intracellular staining profiles of IFNγ, Gata3, phospho-Gata3, T-bet and phospo-Akt1 are shown. The number represents the MFI of the y axis in each area (top). A summary of the MFI in each area (IFNγ: Low, Int, High) is presented (bottom) (n=5). (e) Memory Th2 cells were cultured in IL-7-containing medium with or without Akt inhibitor XI (10 μM) for 2 days. Intracellular staining profiles of IFNγ and IL-4 are shown (left). A summary of the percentage of IL-4- and IFNγ-producing memory Th2 cells is presented (right) (n=4). **P<0.01, *P<0.05 by Student's t-test. Five (a), four (e) and three (b,c,d) independent experiments were performed with similar results.
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f6: Increased phosphorylation status of Gata3 and Akt1 in IFNγ-producing memory Th2 cells.(a) Memory Th2 cells were stimulated with PMA plus ionomycin for 6 h. The cells were divided into four distinct subpopulations according to their expression of IL-4 and IFNγ (left). Intracellular staining profiles of T-bet in these four subpopulations are shown (middle). The number in the histogram represents the MFI. The grey-filled histogram shows isotype control staining. A summary of the MFI of T-bet in IFNγ-producing and -nonproducing cells is presented (right) (n=5). (b) Intracellular staining profiles of IFNγ and IL-4 in effector and memory Th2 cells from WT or Tbx21-deficient mice are shown with the percentages of cells in each area (left). A summary of the percentage of IFNγ-producing memory Th2 cells is presented (right) (n=3). (c) Intracellular staining profiles of Gata3, phospho-Gata3, Akt1 and phospho-Akt1 in IFNγ-producing and -nonproducing memory Th2 cells are shown (left). A summary of the MFI value of Gata3 and phospho-Gata3 and the ratio of the MFI (phospho-Gata3/total Gata3) in IFNγ-producing and IFNγ-nonproducing memory Th2 cells is presented (right) (n=5). (d) Intracellular staining profiles of IFNγ, Gata3, phospho-Gata3, T-bet and phospo-Akt1 are shown. The number represents the MFI of the y axis in each area (top). A summary of the MFI in each area (IFNγ: Low, Int, High) is presented (bottom) (n=5). (e) Memory Th2 cells were cultured in IL-7-containing medium with or without Akt inhibitor XI (10 μM) for 2 days. Intracellular staining profiles of IFNγ and IL-4 are shown (left). A summary of the percentage of IL-4- and IFNγ-producing memory Th2 cells is presented (right) (n=4). **P<0.01, *P<0.05 by Student's t-test. Five (a), four (e) and three (b,c,d) independent experiments were performed with similar results.
Mentions: Recently, distinct memory-type Th2 cell subsets have been identified in several groups, which produce a substantial amount of IL-5, IL-17 or IFNγ in addition to IL-4 and IL-13 (ref. 7). IFNγ expression in Th2 cells is reported to be dependent on T-bet and plays an important role in the protective immunity against viral infections6. We examined whether Gata3 phosphorylation-induced derepression of Tbx21 expression is involved in the IFNγ production from these memory-type Th2 cells. Antigen-specific memory Th2 cells can be efficiently generated in vivo by adoptive transfer of effector Th2 cells34. To determine the expression levels of T-bet in IFNγ-producing memory Th2 cells, the cells were divided into four distinct subpopulations according to their expression of IL-4 and IFNγ. Consistent with previous report35, we found that IFNγ-producing memory Th2 cells express higher levels of T-bet protein as compared with non-IFNγ-producing memory Th2 cells (Fig. 6a). To examine whether IFNγ production from memory Th2 cells is T-bet-dependent, memory Th2 cells from Tbx21-deficient cells were generated. While there was no difference in effector Th2 cell differentiation between WT and Tbx21-deficient cells, IFNγ production from Tbx21-deficient memory Th2 cells was strongly inhibited (Fig. 6b).

Bottom Line: We also identify Akt1 as a Gata3-phosphorylating kinase, and the activation of Akt1 induces derepression of Tbx21 and Ifng expression in Th2 cells.Moreover, T-bet-dependent IFNγ expression in IFNγ-producing memory Th2 cells appears to be controlled by the phosphorylation status of Gata3 in human and murine systems.Thus, this study highlights the molecular basis for posttranslational modifications of Gata3 that control the regulation of IFNγ expression in memory Th2 cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670, Japan.

ABSTRACT
Th2 cells produce Th2 cytokines such as IL-4, IL-5 and IL-13, but repress Th1 cytokine IFNγ. Recent studies have revealed various distinct memory-type Th2 cell subsets, one of which produces a substantial amount of IFNγ in addition to Th2 cytokines, however it remains unclear precisely how these Th2 cells produce IFNγ. We herein show that phosphorylation of Gata3 at Ser308, Thr315 and Ser316 induces dissociation of a histone deacetylase Hdac2 from the Gata3/Chd4 repressive complex in Th2 cells. We also identify Akt1 as a Gata3-phosphorylating kinase, and the activation of Akt1 induces derepression of Tbx21 and Ifng expression in Th2 cells. Moreover, T-bet-dependent IFNγ expression in IFNγ-producing memory Th2 cells appears to be controlled by the phosphorylation status of Gata3 in human and murine systems. Thus, this study highlights the molecular basis for posttranslational modifications of Gata3 that control the regulation of IFNγ expression in memory Th2 cells.

No MeSH data available.


Related in: MedlinePlus