Fig. 5: Upregulation of PI3K/Akt signals in the absence of GFAT1 promotes growth and survival of αβ-T cells while increased ERK signals skew lineage commitment towards γδ-T cells. | Nature Communications

Fig. 5: Upregulation of PI3K/Akt signals in the absence of GFAT1 promotes growth and survival of αβ-T cells while increased ERK signals skew lineage commitment towards γδ-T cells.

From: Dietary glucosamine overcomes the defects in αβ-T cell ontogeny caused by the loss of de novo hexosamine biosynthesis

Fig. 5

Thymocytes from male and female WT, GFAT1- or combined PTEN/GFAT1T−/− mice were used for the following. a–e Thymocytes were stained for Lin, CD4, CD8α, CD25, CD44, CD5,TCRβ, and γδTCR, and intracellularly stained for pS473-Akt (a, b) or pT202/Y204-ERK (c). After flow cytometric analysis, proportion of cells that positively stained for each phosphorylated protein was plotted (a–c). Representative results out of three independent experiments with similar results are shown (a–c). Bar graphs (a, c) represent the amounts (median fluorescence intensity; MFI) of each specific phosphorylation (n = 3–9 mice each). Size (FSC) and granularity (SSC) of DN4 cells expressing TCRβ on their surface and with (pS473+-Akt TCRβ+-DN4) or without phosphorylated Akt at S473 (TCRβ+-DN4 no pS473-Akt) was plotted (b). Numbers in each plot represent proportion of “blasting” cells. Bar graphs (d, e) represent the expression (median fluorescence intensity; MFI) of CD5 (n = 3 mice) (d) as well as either γδTCR or TCRβ (pre-TCR) (n = 6 mice)(e) on the surface of CD4−CD8−Lin− DN thymocytes. Representative FACS plots of the respective receptor staining out of three independent experiments with similar results are shown on the right side of the corresponding bar graph. f, g Thymocytes were surface-stained for γδTCR followed by phospho-ERK intracellular staining. A representative FACS plot of γδTCR staining (f) or phosphorylated-ERK (g) out of three experiments with similar results is shown. Bar graphs represent the median fluorescence intensity (MFI) of γδTCR surface expression (f) or phosphorylated ERK (g) (n = 4 mice each with 3 technical replicates). All bar graphs (a–g) denote mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 using two-sided Student’s t test (c, d) or one-way ANOVA followed by Tukey’s post-hoc test (a, e–g). See also Supplementary Fig 5. Source data are available for (a–g).

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