FIG 9 .

Effect of small TbTim knockdown on import of TbTim17 into mitochondria. (A to D) In vivo import assays for TbTim17-Myc in (A) control, (B) TbTim9 RNAi, (C) TbTim10 RNAi, and (D) TbTim8/13 RNAi T. brucei mitochondria. Mitochondrial proteins were analyzed by Western blots at different time points from h 16 to h 96 using anti-myc and anti-TbTim17 antibodies. TAO was used as a loading control. (E to G) Quantitation of the levels of ectopic TbTim17-Myc using the (E) anti-TbTim17 antibody and (F) anti-Myc antibody and (G) endogenous TbTim17 in all mitochondrial samples during the time course. Intensities of the ectopic TbTim17-Myc and endogenous TbTim17 bands were normalized with the levels of TAO in the corresponding sample. Relative band intensities are presented as percentages of maximum levels of ectopic TbTim17-Myc or endogenous TbTim17 in control mitochondria at 96 h (considered to represent 100%). Error bars indicate standard deviations of results from three independent experiments. (H) Autoradiograph of the in vitro import of radiolabeled TbTim17 in wild-type and small TbTim knockdown mitochondria from min 1 to min 15. Mitochondria were also pretreated with CCCP to disrupt membrane potential (ΔΨ). (I) Quantitation of imported TbTim17 at various time points. The wild-type signal at 15 min was set to 100. Results represent three independent experiments.