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Generation of pathogenic T(H)17 cells in the absence of TGF-β signalling.

Ghoreschi K, Laurence A, Yang XP, Tato CM, McGeachy MJ, Konkel JE, Ramos HL, Wei L, Davidson TS, Bouladoux N, Grainger JR, Chen Q, Kanno Y, Watford WT, Sun HW, Eberl G, Shevach EM, Belkaid Y, Cua DJ, Chen W, O'Shea JJ - Nature (2010)

Bottom Line: Neither IL-6 nor IL-23 alone efficiently generated T(H)17 cells; however, these cytokines in combination with IL-1β effectively induced IL-17 production in naive precursors, independently of TGF-β.T-bet(+)RORγt(+) T(H)17 cells are generated in vivo during experimental allergic encephalomyelitis, and adoptively transferred T(H)17 cells generated with IL-23 without TGF-β1 were pathogenic in this disease model.These data indicate an alternative mode for T(H)17 differentiation.

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunology and Inflammation Branch, National Institute of Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA. ghoreschik@mail.nih.gov

ABSTRACT
CD4(+) T-helper cells that selectively produce interleukin (IL)-17 (T(H)17), are critical for host defence and autoimmunity. Although crucial for T(H)17 cells in vivo, IL-23 has been thought to be incapable of driving initial differentiation. Rather, IL-6 and transforming growth factor (TGF)-β1 have been proposed to be the factors responsible for initiating specification. Here we show that T(H)17 differentiation can occur in the absence of TGF-β signalling. Neither IL-6 nor IL-23 alone efficiently generated T(H)17 cells; however, these cytokines in combination with IL-1β effectively induced IL-17 production in naive precursors, independently of TGF-β. Epigenetic modification of the Il17a, Il17f and Rorc promoters proceeded without TGF-β1, allowing the generation of cells that co-expressed RORγt (encoded by Rorc) and T-bet. T-bet(+)RORγt(+) T(H)17 cells are generated in vivo during experimental allergic encephalomyelitis, and adoptively transferred T(H)17 cells generated with IL-23 without TGF-β1 were pathogenic in this disease model. These data indicate an alternative mode for T(H)17 differentiation. Consistent with genetic data linking IL23R with autoimmunity, our findings re-emphasize the importance of IL-23 and therefore may have therapeutic implications.

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IL-23 upregulates IL-23R and modifies the Il17 and Rorc loci in the absence of TGF-βa, b, Naïve CD4+ T cells were activated in serum-free media without cytokines, with individual cytokines or cytokine combinations as indicated. Il23r expression was analyzed by quantitative RT-PCR (mRNA levels ± s.e.m.) on days 1 to 4 after activation (a) or on day 4 only (b), *P<0.01, **P<0.001. c, IL-6 and IL-23 induce Stat3 binding to Il23r, histone 3 acetylation (H3Ac) and histone 3 lysine 4 trimethylation (H3K4m3) of the Il23r locus as determined by chromatin immunoprecipitation and massive parallel sequencing. d, Naïve CD4+ cells were activated as in (a). Fixed cells were immunoprecipitated with anti-H3K4m3, anti-p300 or anti-H3K9m3 antibodies. Eluted DNA was analyzed by quantitative PCR using primers spanning the promoter regions of Il17a, Il17f and Rorc.
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Figure 2: IL-23 upregulates IL-23R and modifies the Il17 and Rorc loci in the absence of TGF-βa, b, Naïve CD4+ T cells were activated in serum-free media without cytokines, with individual cytokines or cytokine combinations as indicated. Il23r expression was analyzed by quantitative RT-PCR (mRNA levels ± s.e.m.) on days 1 to 4 after activation (a) or on day 4 only (b), *P<0.01, **P<0.001. c, IL-6 and IL-23 induce Stat3 binding to Il23r, histone 3 acetylation (H3Ac) and histone 3 lysine 4 trimethylation (H3K4m3) of the Il23r locus as determined by chromatin immunoprecipitation and massive parallel sequencing. d, Naïve CD4+ cells were activated as in (a). Fixed cells were immunoprecipitated with anti-H3K4m3, anti-p300 or anti-H3K9m3 antibodies. Eluted DNA was analyzed by quantitative PCR using primers spanning the promoter regions of Il17a, Il17f and Rorc.

Mentions: If our model is correct, an essential aspect of Th17 differentiation is the induction of IL-23R independently of TGF-β. We found that IL-6 and IL-1β induced Il23r mRNA expression in the absence of TGF-β. Addition of IL-23 significantly increased expression, whereas TGF-β1 inhibited expression (Fig. 2a, b, Supplementary Fig. 3c). Accordingly, the IL-23-mediated induction of the Th17 phenotype was altered by TGF-β1 (Supplementary Fig. 3d). The inhibitory effect of TGF-β signaling on Il23r expression was also evident using the TGFβRi and in cells with impaired TGF-β signaling (Supplementary Fig. 3e, f). Since IL-23 and IL-6 exert their effects through STAT3, we also documented that STAT3 binds the Il23r locus using chromatin immunoprecipitation with massive parallel sequencing (Fig. 2c).


Generation of pathogenic T(H)17 cells in the absence of TGF-β signalling.

Ghoreschi K, Laurence A, Yang XP, Tato CM, McGeachy MJ, Konkel JE, Ramos HL, Wei L, Davidson TS, Bouladoux N, Grainger JR, Chen Q, Kanno Y, Watford WT, Sun HW, Eberl G, Shevach EM, Belkaid Y, Cua DJ, Chen W, O'Shea JJ - Nature (2010)

IL-23 upregulates IL-23R and modifies the Il17 and Rorc loci in the absence of TGF-βa, b, Naïve CD4+ T cells were activated in serum-free media without cytokines, with individual cytokines or cytokine combinations as indicated. Il23r expression was analyzed by quantitative RT-PCR (mRNA levels ± s.e.m.) on days 1 to 4 after activation (a) or on day 4 only (b), *P<0.01, **P<0.001. c, IL-6 and IL-23 induce Stat3 binding to Il23r, histone 3 acetylation (H3Ac) and histone 3 lysine 4 trimethylation (H3K4m3) of the Il23r locus as determined by chromatin immunoprecipitation and massive parallel sequencing. d, Naïve CD4+ cells were activated as in (a). Fixed cells were immunoprecipitated with anti-H3K4m3, anti-p300 or anti-H3K9m3 antibodies. Eluted DNA was analyzed by quantitative PCR using primers spanning the promoter regions of Il17a, Il17f and Rorc.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3108066&req=5

Figure 2: IL-23 upregulates IL-23R and modifies the Il17 and Rorc loci in the absence of TGF-βa, b, Naïve CD4+ T cells were activated in serum-free media without cytokines, with individual cytokines or cytokine combinations as indicated. Il23r expression was analyzed by quantitative RT-PCR (mRNA levels ± s.e.m.) on days 1 to 4 after activation (a) or on day 4 only (b), *P<0.01, **P<0.001. c, IL-6 and IL-23 induce Stat3 binding to Il23r, histone 3 acetylation (H3Ac) and histone 3 lysine 4 trimethylation (H3K4m3) of the Il23r locus as determined by chromatin immunoprecipitation and massive parallel sequencing. d, Naïve CD4+ cells were activated as in (a). Fixed cells were immunoprecipitated with anti-H3K4m3, anti-p300 or anti-H3K9m3 antibodies. Eluted DNA was analyzed by quantitative PCR using primers spanning the promoter regions of Il17a, Il17f and Rorc.
Mentions: If our model is correct, an essential aspect of Th17 differentiation is the induction of IL-23R independently of TGF-β. We found that IL-6 and IL-1β induced Il23r mRNA expression in the absence of TGF-β. Addition of IL-23 significantly increased expression, whereas TGF-β1 inhibited expression (Fig. 2a, b, Supplementary Fig. 3c). Accordingly, the IL-23-mediated induction of the Th17 phenotype was altered by TGF-β1 (Supplementary Fig. 3d). The inhibitory effect of TGF-β signaling on Il23r expression was also evident using the TGFβRi and in cells with impaired TGF-β signaling (Supplementary Fig. 3e, f). Since IL-23 and IL-6 exert their effects through STAT3, we also documented that STAT3 binds the Il23r locus using chromatin immunoprecipitation with massive parallel sequencing (Fig. 2c).

Bottom Line: Neither IL-6 nor IL-23 alone efficiently generated T(H)17 cells; however, these cytokines in combination with IL-1β effectively induced IL-17 production in naive precursors, independently of TGF-β.T-bet(+)RORγt(+) T(H)17 cells are generated in vivo during experimental allergic encephalomyelitis, and adoptively transferred T(H)17 cells generated with IL-23 without TGF-β1 were pathogenic in this disease model.These data indicate an alternative mode for T(H)17 differentiation.

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunology and Inflammation Branch, National Institute of Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA. ghoreschik@mail.nih.gov

ABSTRACT
CD4(+) T-helper cells that selectively produce interleukin (IL)-17 (T(H)17), are critical for host defence and autoimmunity. Although crucial for T(H)17 cells in vivo, IL-23 has been thought to be incapable of driving initial differentiation. Rather, IL-6 and transforming growth factor (TGF)-β1 have been proposed to be the factors responsible for initiating specification. Here we show that T(H)17 differentiation can occur in the absence of TGF-β signalling. Neither IL-6 nor IL-23 alone efficiently generated T(H)17 cells; however, these cytokines in combination with IL-1β effectively induced IL-17 production in naive precursors, independently of TGF-β. Epigenetic modification of the Il17a, Il17f and Rorc promoters proceeded without TGF-β1, allowing the generation of cells that co-expressed RORγt (encoded by Rorc) and T-bet. T-bet(+)RORγt(+) T(H)17 cells are generated in vivo during experimental allergic encephalomyelitis, and adoptively transferred T(H)17 cells generated with IL-23 without TGF-β1 were pathogenic in this disease model. These data indicate an alternative mode for T(H)17 differentiation. Consistent with genetic data linking IL23R with autoimmunity, our findings re-emphasize the importance of IL-23 and therefore may have therapeutic implications.

Show MeSH
Related in: MedlinePlus