Human urinary bladder epithelial cells lacking wild-type p53 function are deficient in the repair of 4-aminobiphenyl-DNA adducts in genomic DNA.
The effect of the tumor suppressor gene TP53 on repair of genomic DNA damage was examined in human urinary bladder transitional cell carcinoma (TCC) cell lines. Utilizing TCC10 containing wild-type p53 (wt-p53) as the parental line, an isogenic set of cell lines was derived by retroviral infection that expressed a transdominant mutant p53 (Arg --> His at codon 273, TDM273-TCC10), or the human papilloma virus 16-E6 oncoprotein (E6-TCC10). 32P-postlabeling analyses were performed on DNA from TCC cultures obtained after treatment with N-hydroxy-4-aminobiphenyl (N-OH-ABP), N-hydroxy-4-acetylaminobiphenyl (N-OH-AABP) and N-acetoxy-4-acetylaminobiphenyl (N-OAc-AABP). The major adduct was identified as N-(deoxyguanosin-8-yl)-4-aminobiphenyl (dG-C8-ABP) with all three chemicals. The amount of adducts in urothelial DNA ranged between 0.1 and 20 per 10(6) nucleotides, N-OAc-AABP yielding the highest levels, followed by N-OH-ABP and N-OH-AABP. To determine, if the functional status of p53 affects the rate of repair of dG-C8-ABP in genomic DNA, TCC10 and the TDM273-TCC10 and E6-TCC10 isotypes were exposed to N-OH-AABP for 12h and the DNA damage was allowed to repair up to 24h. The adduct levels were quantified and compared between the TCC10 isotypes. The amounts of dG-C8-ABP that remained in genomic DNA from E6-TCC10 and TDM273-TCC10 were approximately two-fold higher, as compared to the parental TCC10. At the dose used for DNA repair studies, N-OH-AABP or N-OAc-AABP did not induce apoptosis in TCC10. However, N-OAc-AABP at high doses (>5 microM) induced apoptosis, as evidenced by DNA fragmentation analyses. Furthermore, N-OAc-AABP-mediated apoptosis was independent of the functional status of wt-p53, since both E6-TCC10 and the parental TCC10 exhibited DNA fragmentation following treatment. These results suggest that p53 might modulate the repair of DNA adducts generated from the human bladder carcinogen ABP in its target human uroepithelial cells. This implies that in p53 null cells the unrepaired DNA damage could cause accumulation of mutation, which might contribute to increased genomic instability and neoplastic progression.
Swaminathan
Santhanam
S
Department of Pharmacology, University of Wisconsin, Madison, WI 53706, USA. swaminat@facstaff.wisc.edu
Torino
Jennifer L
JL
Burger
Melissa S
MS
D000611:Aminobiphenyl Compounds / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D002273:Carcinogens / Q000494:pharmacology; D002478:Cells, Cultured; D003062:Codon; D018736:DNA Adducts / Q000737:chemistry / Q000187:drug effects / Q000502:physiology*; D004260:DNA Repair / Q000187:drug effects / Q000502:physiology; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000473:pathology / Q000502:physiology; D006801:Humans; D009154:Mutation; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000378:metabolism*; D001743:Urinary Bladder / Q000166:cytology / Q000502:physiology*
D000611:Aminobiphenyl Compounds; D002273:Carcinogens; D003062:Codon; D018736:DNA Adducts; C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D012097:Repressor Proteins; D016159:Tumor Suppressor Protein p53; C006757:4-biphenylamine; C022916:N-acetoxy-4-acetylaminobiphenyl; C001365:N-hydroxy-4-acetylaminobiphenyl; C014593:N-hydroxy-4-aminobiphenyl; C053420:N-(deoxyguanosin-8-yl)-4-aminobiphenyl; D003849:Deoxyguanosine