FIGURE 4: The assembly of PulD-FL in mitochondria depends on import receptors. (A) Left panel: Isolated mitochondria were obtained from the indicated strains transformed with either an empty vector (Ø) or a plasmid encoding PulD-FL. Samples were analysed by SDS-PAGE and immunodecoration with the indicated antibodies. Right panel: the steady state levels of PulD-FL in at least three experiments as in the left panel were quantified. The signal of Fis1 was taken as a loading control. Levels of PulD-FL in the corresponding wild type (WT) cells were set to 100%. The bar diagram shows the mean values ± s.d. of at least three independent experiments. (*, P < 0.05; **, P < 0.01; two tailed Student's t-test). (B) Left panel: Crude mitochondria were obtained from WT, tom20μ, or a strain overexpressing TOM20 (Tom20↑) harbouring a plasmid expressing PulD-FL. Samples were analysed by SDS-PAGE and immunodecoration with the indicated antibodies. Right panel: the steady state levels of the PulD-FL secretin were quantified and further analysed as described for part (A). (*, P < 0.05; two-tailed Student's t-test). (C) Crude mitochondria were obtained from WT, tom70/71Δ, or a strain overexpressing TOM70 (Tom70↑) harbouring a plasmid expressing PulD-FL. Further treatment and analysis were as described in the legend of part (B).