Fig. 1.

Decanoic acid causes a reduction in p-4E-BP1 levels in Dictyostelium. (A) Simplified schematic diagram of mTORC1 pathway signaling. Glucose activates mTORC1 through inhibition of AMPK. Conversely, the ketogenic diet inhibits mTORC1 through inhibition of PI3K/AKT signaling. Phosphorylation of 4E-BP1 by mTORC1 induces 4E-BP1 to dissociate from eukaryotic translation initiation factor 4E (eIF4E), promoting the initiation of translation, and this phosphorylation is used in this study as a readout for mTORC1 activation. (B and C) Dictyostelium cells were treated with (B) decanoic acid (n = 12) or (C) octanoic acid (n = 6) for 7 d at a range of concentrations (µM). Percentage growth was plotted normalized to the solvent control (0 µM, 0.2% DMSO) at 7 d. (D) Dose–response curves of normalized growth rate plotted against log concentration of compound were used to calculate IC₅₀ values. (E and F) p-4E-BP1 levels were analyzed in wild-type cells treated for 1 or 24 h with (E) decanoic acid (n = 10) (one-way ANOVA with Dunnett’s post hoc test) or (F) octanoic acid (n = 10) (one-way ANOVA with Dunnett’s post hoc test) at concentrations corresponding to an ∼95% reduction in growth rate. Methylcrotonyl-CoA carboxylase (MCCC1) served as a loading control. (G) AKT activity was evaluated in wild-type cells treated for 1 or 24 h with decanoic acid using an anti–phospho-AKT substrate antibody, with MCCC1 as a loading control (n = 10) (one-way ANOVA with Dunnett’s post hoc test). (H) AMPK activation was analyzed using an antibody against phospho-AMPK, with MCCC1 as a loading control (n = 10) (one-way ANOVA with Dunnett’s post hoc test). Data represent the mean ± SEM. Significance is indicated by ***P ≤ 0.001, ****P ≤ 0.0001.