Ragulator KD increases anterograde lysosome transport. (A) Control-KD, LAMTOR1-KD, or lyspersin-KO cells were allowed to internalize dextran–Alexa Fluor 555 for 6 h at 37°C, chased overnight, and analyzed by live-cell imaging. Images are the first frames from Videos 1, 2, and 3. Bottom panels are magnified views of the boxed areas. Anterograde and retrograde trajectories are represented by red and green lines, respectively. Bars, 5 µm. (B) Long-range lysosome movement was tracked and quantified with ImageJ from control-KD, LAMTOR1-KD, or lyspersin-KO cells (five cells from five independent experiments). Values are the mean ± SD. *, P < 0.05; n.s., not significant (Student’s t test); ^#, P < 0.05; ^##, P < 0.01 (ANOVA). In addition to changes in lysosome positioning and motility, we noticed that LAMTOR1 KD increased the number of lysosomes, probably because of enhanced lysosome biogenesis induced by mTORC1 inactivation and consequent TFEB activation and nuclear translocation (Settembre et al., 2012).