Figure 1
From: L166P mutant DJ-1 promotes cell death by dissociating Bax from mitochondrial Bcl-XL
Subcellular distribution of wild-type DJ-1 and DJ-1(L166P) in mitochondria. (A) HEK293 cells transiently transfected with Myc (control), DJ-1-Myc or DJ-1(L166P)-Myc along with Mito-EGFP were subjected to immunocytochemical staining with anti-Myc antibodies (red) and then labeled with the nuclear marker DAPI (blue). Bar, 10 μm. (B) The percentage of cells transfected with DJ-1-Myc or DJ-1(L166P)-Myc presenting mitochondrial or cytosolic staining in (A) was quantified (mean ± S.E.M., n = 3; by one-way ANOVA). (C) H1299 cells transiently transfected with Flag, Flag-DJ-1 or Flag-DJ-1(L166P) were subjected to subcellular fractionation. The total cell lysate and the cytosolic and mitochondrial fractions were subjected to immunoblot analysis. (D) The distribution ratio of Flag-DJ-1 or Flag-DJ-1(L166P) in cytosolic or mitochondrial fractions to their total level in (C) were analyzed using densitometric analysis (mean ± S.E.M., n = 3; by one-way ANOVA). (E) H1299 cells transiently transfected with Flag-DJ-1(L166P) were treated with or without 80 mJ/cm2 UVB irradiation, and the cells were then subjected to the subcellular fractionation assay. The total cell lysate, cytosolic and mitochondrial fractions were subjected to immunoblot analysis. (F) The relative level of Flag-DJ-1(L166P) in (E) were analyzed using densitometric analysis (mean ± S.E.M., n = 3; by one-way ANOVA).