Figure 1

Only paclitaxel causes accumulation of punctate foci of GFP-LC3 in mitotic cells. (A) Accumulation of GFP-LC3 punctate foci in cells treated with either paclitaxel or nocodazole overnight. Mitochondria are shown in red after exposure to MitoTracker Red CMXRos dye [32, 39]. The punctate foci of bright green GFP-LC3 in stably transfected HeLa cells overlapping with weak red punctate foci of dysfunctional mitochondria are indicative of mitophagosomes or mitolysosomes (mitochondria-containing autolysosomes). Colocalized punctate foci in white were revealed by analysis with an ImageJ ColocalizeRGB Plugin. The GFP-LC3 punctate foci overlapping with the much weaker signal of MitoTracker were not detected in the colocalization analysis. Lysosomal inhibitor NH₄Cl (20 mM NH4Cl) was added simultaneously to preserve the resulting mitophagosomes. Scale bar, 10 μm in all panels. Ctrl, Control, Noc, 10 μM nocodazole; Tax, 10 μM paclitaxel. (B) Fluorescent intensity of GFP-LC3 in mitotic cells treated with paclitaxel and nocodazole overnight. (C) The percentage of pro-metaphase cells with punctate foci of GFP-LC3 as shown for the mitotic cells treated with paclitaxel overnight in the absence of NH₄Cl (A). Number of cells with GFP-LC3 punctate foci was counted directly and expressed as a percentage of total mitotic cells. Because of the difference in frequency of mitosis, about 100 mitotic cells in 20 to 50 different fields were scored in the presence or absence of nocodazole or paclitaxel, respectively. The data represent the means and standard errors of three independent experiments. **, p < 0.01 as calculated by Student's T-test.