Figure 7.

F-Actin in Mitochondria, as Indicated by Staining with the F-Actin–Specific Fluorescent Dye Phallacidin.

(A) Isolated mitochondria washed briefly with 1 M KCl followed by various concentrations of Lat B treatment with or without 5 μM phalloidin pretreatment. Flow cytometric analysis indicates that KCl stripped mitochondria can be stained by the F-actin–specific dye FL phallacidin (red curve) and that this fluorescence is reduced by treatment with the F-actin depolymerization reagent Lat B (20 and 60 μM, respectively, shown in the green and blue curves). The stabilization of mitochondrial F-actin by phalloidin (5 μg mL⁻¹) rescues the effect of Lat B on mitochondrial F-actin and retains the mitochondrial phallacidin fluorescence intensity (brown and black curves) in flow cytometric analyses. For each experiment, 500,000 mitochondria were analyzed.

(B) Fluorescence images indicate that the fluorescence of mitochondrial filamentous actin stained by fluorescent phallacidin was decreased caused by Lat B (20 and 60 μM) treatment. This result provides the same conclusion as that in (A) and indicates that actin in mitochondria exists in a filamentous form and that Lat B treatment decreases the fluorescence of FL phallacidin-stained mitochondria. Differential interference contrast (DIC) images in the bottom row represent individual mitochondrion existing in each image of the top row correspondingly.