Fig. 4
2-Undecanone alters sphingomyelin metabolism and destabilizes lysosomes. (A) Acid sphingomyelinase (ASM) activity in lysosomes. WT (2-U), lysosomes from 2-undecanone-treated worms. WT (Control), lysosomes from 2-undecanone-control worms. hsp-1 (tm5076), lysosomes from non-treatment hsp-1 worms. WT, lysosomes from non-treatment worms. Quantitative PCR (qPCR) analysis of asm-1, asm-2, asm-3 (B), asah-1, asah-2 (C) genes levels in 2-undecanone-treated C. elegans and 2-undecanone-control C. elegans for 3 h. Cell division control protein 42 homolog (cdc-42) served as an internal control. (D) Sensitivity of lysosomes of asm-1, asm-2, asm-3 mutants stained by intestinal lysosomal marker Lyso-Tracker to 2-undecanone. Scale bars, 20 μm. Expression levels of ASM-1, ASM-2 and ASM-3 of 2-undecanone-treated worms and hsp-1 (tm5076) mutants in lysosomes with LMP-1 as an internal control (E). Total protein with β-Actin as an internal control (F). In total protein, β-Actin served as an internal control. (G) SM concentrations of lysosomes as measured by Sphingomyelin assay kit. (H) Ceramide concentration of lysosomes was measured based on HPLC- fluorescence detecting method. The data represents means ± SD from triplicate experiments. *P < 0.05, **P < 0.01, ***P < 0.001.
