FIGURE 5.

H₂O₂ induction of 8-OHdG in DNA of Fhit and Fhit mutant stable and induced expressors. A, stable MKN74 Fhit and mutant expressors were treated with 0.5 mm H₂O₂, and DNA damage in the nuclear genomes was assessed by immunofluorescent detection of 8-OHdG residues in the DNA of the respective cell lines. The red nuclear stain detects 8-OHdG and the blue stain is 4′,6-diamidino-2-phenylindole nuclear DNA staining (insets). Negative controls for all cells showed no staining (data not shown). B, bar graph ± S.D. shows the relative frequency of 8-OHdG detection in MKN74 WT and mutant expressors. The double asterisk on Y114A and H96D denotes a p value >0.05. C, immunoblot analysis of Fdxr, Fhit, and glyceraldehyde-3-phosphate dehydrogenase in H1299 cells expressing Fhit WT, Y114A, and Y114D proteins, showing the Fdxr level after cycloheximide (CHX) chase (30 μg/ml) for 6–24 h. Densitometry based on glyceraldehyde-3-phosphate dehydrogenase levels shows enhanced stability of Fdxr in presence of WT Fhit.