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Ndfip-mediated degradation of Jak1 tunes cytokine signalling to limit expansion of CD4+ effector T cells.

O'Leary CE, Riling CR, Spruce LA, Ding H, Kumar S, Deng G, Liu Y, Seeholzer SH, Oliver PM - Nat Commun (2016)

Bottom Line: Nedd4 family E3 ubiquitin ligases have been shown to restrict T-cell function and impact T-cell differentiation.We show here that Ndfip1 and Ndfip2, activators of Nedd4 family ligases, together limit accumulation and function of effector CD4+ T cells.In wild-type cells, Jak1 degradation lessens CD4+ cell sensitivity to cytokines during TCR stimulation, while in Ndfip-deficient cells cytokine responsiveness persists, promoting increased expansion and survival of pathogenic effector T cells.

View Article: PubMed Central - PubMed

Affiliation: Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

ABSTRACT
Nedd4 family E3 ubiquitin ligases have been shown to restrict T-cell function and impact T-cell differentiation. We show here that Ndfip1 and Ndfip2, activators of Nedd4 family ligases, together limit accumulation and function of effector CD4+ T cells. Using a three-part proteomics approach in primary T cells, we identify stabilization of Jak1 in Ndfip1/2-deficient T cells stimulated through the TCR. Jak1 degradation is aborted in activated T cells that lack Ndfips. In wild-type cells, Jak1 degradation lessens CD4+ cell sensitivity to cytokines during TCR stimulation, while in Ndfip-deficient cells cytokine responsiveness persists, promoting increased expansion and survival of pathogenic effector T cells. Thus, Ndfip1/Ndfip2 regulate the cross talk between the T-cell receptor and cytokine signalling pathways to limit inappropriate T-cell responses.

No MeSH data available.


Related in: MedlinePlus

Ndfip2−/− mice do not show signs of inflammation.(a,b) Representative flow cytometry analysis of T-cell populations from thymus and spleen of 5–7-week-old Ndfip2−/− and age-matched control mice: (a) CD4+ and CD8+ cells, (b) CD44 and CD62L expression on these cells, as noted. (c) Intracellular cytokine staining for IL-4 and IFNγ in CD4+ T cells from Ndfip2−/− and WT spleens stimulated ex vivo with PMA and ionomycin in the presence of BFA. Representative of at least five mice per genotype, 5–7 weeks of age. (d) CD4+ T cells were stimulated in vitro for the indicated time periods with αCD3/CD28. Ndfip1 and Ndfip2 expression was analysed by qPCR. Ndfip1/Ndfip2 expression relative to Actb was normalized to expression in unstimulated CD4+ T cells. Representative of a minimum of three independent experiments.
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f1: Ndfip2−/− mice do not show signs of inflammation.(a,b) Representative flow cytometry analysis of T-cell populations from thymus and spleen of 5–7-week-old Ndfip2−/− and age-matched control mice: (a) CD4+ and CD8+ cells, (b) CD44 and CD62L expression on these cells, as noted. (c) Intracellular cytokine staining for IL-4 and IFNγ in CD4+ T cells from Ndfip2−/− and WT spleens stimulated ex vivo with PMA and ionomycin in the presence of BFA. Representative of at least five mice per genotype, 5–7 weeks of age. (d) CD4+ T cells were stimulated in vitro for the indicated time periods with αCD3/CD28. Ndfip1 and Ndfip2 expression was analysed by qPCR. Ndfip1/Ndfip2 expression relative to Actb was normalized to expression in unstimulated CD4+ T cells. Representative of a minimum of three independent experiments.

Mentions: As our GFP reporter indicated high Ndfip2 expression in T cells, we focused our analysis of Ndfip2−/− mice on the T-cell compartment. Ndfip2−/− mice had normal thymic populations and normal CD4+ and CD8+ T cell percentages in lymph nodes and spleens (Fig. 1a; Supplementary Fig. 3a). Ndfip2−/− mice had no increase in percent of CD44+ T cells or cytokine production upon ex vivo analysis (Fig. 1b,c; Supplementary Fig. 3a). Helper T-cell differentiation in vitro, measured by expression of lineage defining transcription factors and cytokine production, was similar between Ndifp2−/− and control cells (Supplementary Fig. 3b,c).


Ndfip-mediated degradation of Jak1 tunes cytokine signalling to limit expansion of CD4+ effector T cells.

O'Leary CE, Riling CR, Spruce LA, Ding H, Kumar S, Deng G, Liu Y, Seeholzer SH, Oliver PM - Nat Commun (2016)

Ndfip2−/− mice do not show signs of inflammation.(a,b) Representative flow cytometry analysis of T-cell populations from thymus and spleen of 5–7-week-old Ndfip2−/− and age-matched control mice: (a) CD4+ and CD8+ cells, (b) CD44 and CD62L expression on these cells, as noted. (c) Intracellular cytokine staining for IL-4 and IFNγ in CD4+ T cells from Ndfip2−/− and WT spleens stimulated ex vivo with PMA and ionomycin in the presence of BFA. Representative of at least five mice per genotype, 5–7 weeks of age. (d) CD4+ T cells were stimulated in vitro for the indicated time periods with αCD3/CD28. Ndfip1 and Ndfip2 expression was analysed by qPCR. Ndfip1/Ndfip2 expression relative to Actb was normalized to expression in unstimulated CD4+ T cells. Representative of a minimum of three independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4837450&req=5

f1: Ndfip2−/− mice do not show signs of inflammation.(a,b) Representative flow cytometry analysis of T-cell populations from thymus and spleen of 5–7-week-old Ndfip2−/− and age-matched control mice: (a) CD4+ and CD8+ cells, (b) CD44 and CD62L expression on these cells, as noted. (c) Intracellular cytokine staining for IL-4 and IFNγ in CD4+ T cells from Ndfip2−/− and WT spleens stimulated ex vivo with PMA and ionomycin in the presence of BFA. Representative of at least five mice per genotype, 5–7 weeks of age. (d) CD4+ T cells were stimulated in vitro for the indicated time periods with αCD3/CD28. Ndfip1 and Ndfip2 expression was analysed by qPCR. Ndfip1/Ndfip2 expression relative to Actb was normalized to expression in unstimulated CD4+ T cells. Representative of a minimum of three independent experiments.
Mentions: As our GFP reporter indicated high Ndfip2 expression in T cells, we focused our analysis of Ndfip2−/− mice on the T-cell compartment. Ndfip2−/− mice had normal thymic populations and normal CD4+ and CD8+ T cell percentages in lymph nodes and spleens (Fig. 1a; Supplementary Fig. 3a). Ndfip2−/− mice had no increase in percent of CD44+ T cells or cytokine production upon ex vivo analysis (Fig. 1b,c; Supplementary Fig. 3a). Helper T-cell differentiation in vitro, measured by expression of lineage defining transcription factors and cytokine production, was similar between Ndifp2−/− and control cells (Supplementary Fig. 3b,c).

Bottom Line: Nedd4 family E3 ubiquitin ligases have been shown to restrict T-cell function and impact T-cell differentiation.We show here that Ndfip1 and Ndfip2, activators of Nedd4 family ligases, together limit accumulation and function of effector CD4+ T cells.In wild-type cells, Jak1 degradation lessens CD4+ cell sensitivity to cytokines during TCR stimulation, while in Ndfip-deficient cells cytokine responsiveness persists, promoting increased expansion and survival of pathogenic effector T cells.

View Article: PubMed Central - PubMed

Affiliation: Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

ABSTRACT
Nedd4 family E3 ubiquitin ligases have been shown to restrict T-cell function and impact T-cell differentiation. We show here that Ndfip1 and Ndfip2, activators of Nedd4 family ligases, together limit accumulation and function of effector CD4+ T cells. Using a three-part proteomics approach in primary T cells, we identify stabilization of Jak1 in Ndfip1/2-deficient T cells stimulated through the TCR. Jak1 degradation is aborted in activated T cells that lack Ndfips. In wild-type cells, Jak1 degradation lessens CD4+ cell sensitivity to cytokines during TCR stimulation, while in Ndfip-deficient cells cytokine responsiveness persists, promoting increased expansion and survival of pathogenic effector T cells. Thus, Ndfip1/Ndfip2 regulate the cross talk between the T-cell receptor and cytokine signalling pathways to limit inappropriate T-cell responses.

No MeSH data available.


Related in: MedlinePlus