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. 2013 Oct 3;9(10):e1003857. doi: 10.1371/journal.pgen.1003857

Figure 4. ESCO2 mutation is associated with mTOR inhibition in RBS cells.

Figure 4

(A–D). Phosphorylation of S6K1/S6 and 4EBP1-γ subunit ratio divided by α/β subunit was downregulated in the human immortalized RBS cells (A–B) and primary cells (C–D), as measured by Western blot analysis. The results are representative of three independent experiments. Total levels of S6, S6K1, and tubulin serve as loading controls. The relative ratios of protein expression were quantitated with ImageQuant TL software in (B, D). (E). L-Leu application partially improved poor proliferation of immortalized RBS cells. Each plot represents the average ± SEM of the ratio of the measurement of the indicated cell number, as calculated for three independent samples for 2-way ANOVA statistical analysis. P<0.01, HSF ESCO2-Mut with 10 mM D-Leu vs HSF ESCO2-WT with10 mM D-Leu; P<0.05, HSF ESCO2-Mut with 10 mM D-Leu vs HSF ESCO2-Mut with 10 mM L-Leu. (F). Elevated levels of cell death were partially suppressed by L-Leu supplementation. P<0.01, HSF ESCO2-Mut with 10 mM D-Leu vs HSF ESCO2-WT with 10 mM D-Leu; P<0.05, HSF ESCO2-Mut with 10 mM L-Leu vs HSF ESCO2-Mut with 10 mM D-Leu. (G). L-Leu, but not D-Leu, partially rescued the phosphorylated form of S6 in primary RBS fibroblasts. p53 levels were not rescued by treatment with L-Leu.