Genetic Effects of Oxidative DNA Damages: Comparative Mutagenesis of the Imidazole Ring-Opened Formamidopyrimidines Fapy Lesions and 8-Oxo-Purines in Simian Kidney CellsReportar como inadecuado


Genetic Effects of Oxidative DNA Damages: Comparative Mutagenesis of the Imidazole Ring-Opened Formamidopyrimidines Fapy Lesions and 8-Oxo-Purines in Simian Kidney Cells


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Citation

Kalam, M. Abul, Kazuhiro Haraguchi, Sushil Chandani, Edward L. Loechler, Maasaki Moriya, Marc M. Greenberg, Ashis K. Basu. -Genetic effects of oxidative DNA damages: comparative mutagenesis of the imidazole ring-opened formamidopyrimidines Fapy lesions and 8-oxo-purines in simian kidney cells- Nucleic Acids Research 348: 2305-2315. 2006

Abstract

Fapy·dG and 8-oxo-7,8-dihydro-2′-deoxyguanosine 8-oxo-dG are formed in DNA by hydroxyl radical damage. In order to study replication past these lesions in cells, we constructed a single-stranded shuttle vector containing the lesion in 5′-TGT and 5′-TGA sequence contexts. Replication of the modified vector in simian kidney COS-7 cells showed that Fapy·dG is mutagenic inducing primarily targeted Fapy·G→T transversions. In the 5′-TGT sequence mutational frequency of Fapy·dG was ^∼30%, whereas in the 5′-TGA sequence it was ^∼8%. In parallel studies 8-oxo-dG was found to be slightly less mutagenic than Fapy·dG, though it also exhibited a similar context effect: 4-fold G→T transversions 24% versus 6% occurred in the 5′-TGT sequence relative to 5′-TGA. To investigate a possible structural basis for the higher G→T mutations induced by both lesions when their 3′ neighbor was T, we carried out a molecular modeling investigation in the active site of DNA polymerase β, which is known to incorporate both dCTP no mutation and dATP G→T substitution opposite 8-oxo-G. In pol β, the syn-8-oxo-G:dATP pair showed greater stacking with the 3′-T:A base pair in the 5′-TGT sequence compared with the 3′-A:T in the 5′-TGA sequence, whereas stacking for the anti-8-oxo-G:dCTP pair was similar in both 5′-TGT and 5′-TGA sequences. Similarly, syn-Fapy·G:dATP pairing showed greater stacking in the 5′-TGT sequence compared with the 5′-TGA sequence, while stacking for anti-Fapy·G:dCTP pairs was similar in the two sequences. Thus, for both lesions less efficient base stacking between the lesion:dATP pair and the 3′-A:T base pair in the 5′-TGA sequence might cause lower G→T mutational frequencies in the 5′-TGA sequence compared to 5′-TGT. The corresponding lesions derived from 2′-deoxyadenosine, Fapy·dA and 8-oxo-dA, were not detectably mutagenic in the 5′-TAT sequence, and were only weakly mutagenic <1% in the 5′-TAA sequence context, where both lesions induced targeted A→C transversions. To our knowledge this is the first investigation using extrachromosomal probes containing a Fapy·dG or Fapy·dA site-specifically incorporated, which showed unequivocally that in simian kidney cells Fapy·G→T substitutions occur at a higher frequency than 8-oxo-G→T and that Fapy·dA is very weakly mutagenic, as is 8-oxo-dA.

CAS: Biology: Scholarly Papers -



Autor: Kalam, M. Abul - Haraguchi, Kazuhiro - Chandani, Sushil - Loechler, Edward L. - Moriya, Maasaki - Greenberg, Marc M. - Basu, Ashi

Fuente: https://open.bu.edu/







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