FIG 1.

L. amazonensis APX is a mitochondrial protein upregulated by ROS. (A) APX localizes to mitochondria. Immunolocalization of APX in L. amazonensis promastigotes was performed using antibodies against APX (a [green]). Nucleus and kinetoplast DNA was stained with DAPI (b [blue]), and mitochondria were stained with MitoTracker Red (c [red]). Merging the two images (d [merge]) confirmed the mitochondrial localization of APX (Pearson’s correlation coefficients were 0.826, 0.773, 0.49, and 0.641 in cells 1 to 4). Bar = 4 μm. The extent of overlap between APX staining (green) and mitochondrial staining (red) was determined for individual cells. (B) APX expression is induced by ROS exposure. Log-phase L. amazonensis promastigotes (2 × 10⁷/ml) were treated for 8 h with increasing concentrations of H₂O₂ or menadione, as indicated. Western blots of whole-cell lysates (10 μg protein/lane) were used to compare the amounts of APX protein in different ROS-treated samples. Tubulin expression was used as a loading control. The ratio of APX relative to tubulin in each sample is indicated below each lane and values represent the mean ± SD of three independent experiments.