UV-inducible base excision repair of oxidative damaged DNA in human cells.
Methylene blue (MB) acts as a photosensitizer and after excitation by visible light (VL) produces reactive oxygen species that result in oxidatively damaged DNA. (MB + VL) produces predominantly 8-hydroxyguanine as well as other single base modifications in DNA that are repaired by base excision repair (BER). We have used a recombinant non-replicating human adenovirus, Ad5HCMVlacZ, which expresses the beta-galactosidase (beta-gal) reporter gene, to examine the role of the p53 tumor suppressor in constitutive and inducible BER of MB + VL-damaged DNA in human cells. Host cell reactivation (HCR) of beta-gal activity for MB + VL-treated Ad5HCMVlacZ was examined in normal human fibroblasts and several transformed and tumor cell lines with compromised p53 function using both non-treated cells and cells pretreated with ultraviolet light of 200-280 nm wavelength (UVC). Constitutive HCR of the MB + VL-treated reporter gene in untreated cells did not correlate with wild-type p53 expression levels, suggesting that factors other than p53 expression levels can influence constitutive BER of the reporter gene. UVC pre-treatment of the normal fibroblast strains resulted in an enhanced HCR of the MB + VL-treated reporter gene and a concomitant increase in the expression of p53, suggesting that p53 may be involved in UV-inducible BER in normal human fibroblasts. In contrast, p53 expression did not correlate with HCR values for the p53-compromised cells in UVC-pre-treated cells. In particular, the SKOV-3, LFS 087 and NF-E6 cells showed no up-regulation of p53 expression following UVC, and yet these cells showed significant enhancement of HCR following UVC pre-treatment. These results indicate that BER of MB + VL-damaged DNA is inducible in human cells by pre-UVC treatment and that the enhancement in BER may result from both p53-dependent and p53-independent mechanisms.
Kassam
Shaqil N
SN
Department of Biology, McMaster University, Ontario, Canada.
Rainbow
Andrew J
AJ
D002460:Cell Line; D004247:DNA / Q000528:radiation effects*; D004249:DNA Damage / Q000235:genetics*; D004260:DNA Repair; D006801:Humans; D008751:Methylene Blue / Q000528:radiation effects / Q000633:toxicity; D018384:Oxidative Stress / Q000235:genetics*; D011838:Radiation-Sensitizing Agents / Q000633:toxicity; D017382:Reactive Oxygen Species / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis; D014466:Ultraviolet Rays / Q000009:adverse effects*; D001616:beta-Galactosidase / Q000235:genetics
D011838:Radiation-Sensitizing Agents; D017382:Reactive Oxygen Species; D016159:Tumor Suppressor Protein p53; D004247:DNA; D001616:beta-Galactosidase; D008751:Methylene Blue