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. 2018 Jun 25;131(12):jcs213314. doi: 10.1242/jcs.213314

Fig. 5.

Fig. 5.

Endogenous and synthetic sphingolipids that downregulate nutrient transporters also inactivate the ARF6 GTPase. (A) ARF6-GTP levels normalized to total ARF6 in FL5.12 cells treated with the cytohesin inhibitor SecinH3 (30 µM), C2-ceramide (50 µM), bacterial sphingomyelinase (bSMase; 100 mU/ml), FTY720 (5 µM) or 893 (5 µM) for 3 h. (B,C) ARF6-GTP levels in FL5.12 cells treated with FTY720 (0, 1.25, 2.5, 5 or 10 µM) for 30 min (B) or with 10 µM FTY720 for 5, 15 or 30 min (C). (D) ARF6-GTP levels in FL5.12, MDA-MB-231, HeLa, SW620, PC3 or SupB15 treated for 3 h with FTY720 (10 µM). (E) Steady state surface CD98 levels in FL5.12 cells treated with C2-ceramide or C2-dihydroceramide at the indicated concentrations for 3 h. (F) Viability of C2-ceramide- or C2-dihydroceramide-treated FL5.12 cells at 48 h. (G) ARF6-GTP levels in FL5.12 cells treated with C2-ceramide (50 µM) or C2-dihydroceramide (50 µM). (H) ARF6-GTP levels in FL5.12 cells treated with CalyculinA (10 nM) for 1 h. Means±s.e.m. shown of n≥3 in all panels. Using two-tailed t-test (D-F,H) or ordinary one-way ANOVA (A-C,G): n.s., not significant; *P≤0.05; **P≤0.01; ***P≤0.001. Dunnett's test was used to correct for multiple comparisons (A-C,G). See also Fig. S5.