Figure 5
Asteltoxin controls the number of lysosomes and multivesicular bodies. (A) Electron microscopy images of lysosomes (red arrowheads) in PC3 cells treated with DMSO or asteltoxin (10 μg/mL) and DMSO for 24 h. Scale bar = 10 μm (upper panels) and scale bar = 1 μm (lower panels). (B) Quantification of the number of lysosomes per cell (n = 11 for DMSO and n = 11 for asteltoxin). MVB-lysosome fusion was counted as lysosome. Boxes represent the interquartile range, and the line inside the box represents the median value. (C) Electron microscopy images of MVBs (blue arrowheads) and MVB–lysosome fusion (green arrowheads) in the cells used in (A). Scale bar = 2 μm (upper panels) and scale bar = 1 μm (lower panels). (D) Quantification of the number of MVBs in the fields (n = 50 for DMSO and n = 50 for asteltoxin). The median and interquartile range are shown by bars. (E) Schematic model of the role that asteltoxin plays in regulating the fate of multivesicular bodies (MVBs) and controlling EV secretion. (a) In cancer cells, MVBs fuse with the plasma membrane and promote EV secretion. (b) After treatment with asteltoxin at low concentrations, the AMP/ATP ratio increases due to inhibition of ATP synthase, and this increase induces AMPK-mediated suppression of mTORC1. Inactivation of mTORC1 promotes nuclear translocation of MiT/TFE family members, thereby inducing transcription of lysosome-related genes and activation of lysosomal function. MVBs are then degraded, and the number of MVBs fused with the plasma membrane is low, resulting in suppression of EV secretion. Statistical analysis was performed using one-way ANOVA. *P < 0.05 and **P < 0.01.
