Fig. 6. PEX19 is required for the targeting of PEX26. (A) PEX26 localizes to mitochondria in PEX19-deficient cells. A GFP-PEX26_2-305-expressing plasmid was transfected into PEX19-deficient human fibroblasts and the localization of the GFP fusion protein was determined. Mitochondria were labelled by indirect immunofluorescence using anti-TRAP1 antibodies (Mc). (B) Expression of PEX19 is downregulated in human fibroblasts treated with PEX19 siRNA. Wild-type fibroblasts were transfected with PEX19-specific siRNA or with PEX5 control siRNA and incubated for 5 days at 37°C. Samples were taken every 24 hours and whole-cell lysates were prepared. Equal amounts of protein were analyzed by immunoblotting using antibodies against PEX19 and PEX14. (C,D) Inhibition of PEX19 affects targeting of PEX26. After 60 hours, PEX19 siRNA-treated cells were transfected with a GFP-PEX26_2-305-expressing plasmid. 24 hours later, cells were analyzed for the distribution of the fusion protein. Peroxisomes (Px) and mitochondria (Mc) were visualized by indirect immunofluorescence using anti-PEX14 (C) and anti-TRAP1 (D) antibodies, respectively. Bar, 10 µm.