DNA damage and nuclear translocation of parkin. (A) DNA induces nuclear translocation of parkin. SH-SY5Y cells stably transfected with parkin were UV-irradiated (60 J/m2), or treated with H2O2 (300 μM) or etoposide (50 μM) to induce DNA damage, followed by the isolation of nuclear and cytoplasmic fractions and resolution of parkin by immunoblotting. Note that each DNA damaging agent induced an increase in the absolute and relative levels of parkin in the nucleus. Absence of cross-contamination of the fractions was confirmed by immunoblotting for lamin B, a nuclear marker, and β-actin, a cytoplasmic marker. Quantitation of the relative nuclear ratio of parkin is shown, and represents the mean ± S.D., n = 3. *P < 0.05 relative to control by ANOVA with post-hoc Student Neumann-Kiels tests. (B) Nuclear translocation of parkin in SH-SY5Y cells was confirmed by immunofluorescence assays. SH-SY5Y cells stably over-expressing parkin were UV-irradiated (60 J/m2) or mock-treated. Two hours later the cellular localization of parkin was determined by immunofluorescent microscopy. Green: parkin. Blue: Hoechst staining of nucleus.