Mdm2 as a sensitive and mechanistically informative marker for genotoxicity induced by benzo[a]pyrene and dibenzo[a,l]pyrene.
Mdm2 is an oncoprotein interacting with p53 and maintaining low p53 levels in unstressed cells. Here we investigated the effect of genotoxic compounds on Mdm2 phosphorylation levels. Employing the Mdm2 2A10 antibody and phosphatase treatment we found that Mdm2 accumulated in HepG2 cells when exposed to low concentrations of genotoxic compounds such as mitomycin C, etoposide, 5-fluorouracil, and benzo[a]pyrene (BP). The low-dose responses were not accompanied by p53 accumulation and the effect of low concentrations of BP on Mdm2 was not affected by small interfering RNA for p53. In human lymphoblasts 10nM BP induced an Mdm2 response. Low concentrations of BP also induced binding of Mdm2 to chromatin in HepG2 cells, but no p53 binding or H2AX phosphorylation. The more mutagenic dibenzo[a,l]pyrene as well as higher BP concentrations instead induced gammaH2AX and p53 Ser15 association with chromatin. Acrolein potentiated the effect of BP on p53 stabilization and chromatin binding. Taken together, these data suggest that (1) Mdm2 is a sensitive biomarker for certain types of genotoxicity, and (2) that polycyclic aromatic hydrocarbons-induced Mdm2 binding to chromatin reflects repairable damage, whereas chromatin binding of p53 Ser15 and gammaH2AX indicates more persistent DNA damage. The analysis of Mdm2 and related endpoints might be useful for evaluating mutagenic potentials of DNA damages. It is suggested that patterns documented here can be used for separating BP doses that induce readily repaired DNA adducts from doses that overwhelm this capacity.
Malmlöf
Maria
M
Institute of Environmental Medicine, Karolinska Institutet, S-17177 Stockholm, Sweden.
Pääjärvi
Gerd
G
Högberg
Johan
J
Stenius
Ulla
U
D001564:Benzo(a)pyrene / Q000633:toxicity*; D001580:Benzopyrenes / Q000633:toxicity*; D002273:Carcinogens / Q000633:toxicity*; D045744:Cell Line, Tumor; D002843:Chromatin / Q000187:drug effects / Q000378:metabolism; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000032:analysis / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D005819:Genetic Markers; D006801:Humans; D009153:Mutagens / Q000633:toxicity*; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D051736:Proto-Oncogene Proteins c-mdm2 / Q000235:genetics* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D014162:Transfection
D001580:Benzopyrenes; D002273:Carcinogens; D002843:Chromatin; D004273:DNA, Neoplasm; D005819:Genetic Markers; D009153:Mutagens; D034741:RNA, Small Interfering; D001564:Benzo(a)pyrene; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; C041517:dibenzo(a,l)pyrene