TY - JOUR
T1 - Human DNA polymerases catalyze lesion bypass across benzo[a]pyrene-derived DNA adduct clustered with an abasic site
AU - Starostenko, Lidia V.
AU - Rechkunova, Nadejda I.
AU - Lebedeva, Natalia A.
AU - Kolbanovskiy, Alexander
AU - Geacintov, Nicholas E.
AU - Lavrik, Olga I.
N1 - Funding Information:
The authors are grateful to Dr. R.G. Woodgate and Dr. Ekaterina Frank for the recombinant plasmid DNA coding hPolι and producing strain, Dr. S.H. Wilson for the recombinant plasmid DNA coding hPolλ, Dr. S.N. Khodyreva for the recombinant purified UDG and APE1, Natalia Gayko for technical assistance. This work was partly supported by RFBR grant no. 14-04-00268 (O.L.), by RSCF grant no. 14-14-00501 (L.S., N.R., and N.L.), by grant from the Russian Ministry of Education and Science for Leading Scientific Schools (420.2014.4), and by the Program of the RAS Presidium on Molecular and Cell Biology.
Publisher Copyright:
© 2014 Elsevier B.V.
PY - 2014/12/1
Y1 - 2014/12/1
N2 - The combined action of oxidative stress and genotoxic polycyclic aromatic hydrocarbons derivatives can lead to cluster-type DNA damage that includes both a modified nucleotide and a bulky lesion. As an example, we investigated the possibility of repair of an AP site located opposite a minor groove-positioned (+)-trans-BPDE-dG or a base-displaced intercalated (+)-cis-BPDE-dG adduct (BP lesion) by a BER system. Oligonucleotides with single uracil residue in the certain position were annealed with complementary oligonucleotides bearing either a cis- or trans-BP adduct. Digestion with uracil DNA glycosylase was utilized to generate an AP site which was then hydrolyzed by APE1, and the resulting gap was processed by X-family DNA polymerases β (Polβ) and λ (Polλ), or Y-family polymerase β (Polβ). By varying reaction conditions, namely, Mg2+/Mn2+ replacement/combination and ionic strength decrease, we found that under certain conditions both Polβ and Polι can catalyze lesion bypass across both cis- and trans-BP adducts in the presence of physiological dNTP concentrations. Polβ and Polι catalyze gap filling trans-lesion synthesis in an error prone manner. By contrast, Polλ selectively introduced the correct dCTP opposite the modified dG in the case of cis-BP-dG adduct only, and did not bypass the stereoisomeric trans-adduct under any of the conditions examined. The results suggest that Polλ is a specialized polymerase that can process these kinds of lesions.
AB - The combined action of oxidative stress and genotoxic polycyclic aromatic hydrocarbons derivatives can lead to cluster-type DNA damage that includes both a modified nucleotide and a bulky lesion. As an example, we investigated the possibility of repair of an AP site located opposite a minor groove-positioned (+)-trans-BPDE-dG or a base-displaced intercalated (+)-cis-BPDE-dG adduct (BP lesion) by a BER system. Oligonucleotides with single uracil residue in the certain position were annealed with complementary oligonucleotides bearing either a cis- or trans-BP adduct. Digestion with uracil DNA glycosylase was utilized to generate an AP site which was then hydrolyzed by APE1, and the resulting gap was processed by X-family DNA polymerases β (Polβ) and λ (Polλ), or Y-family polymerase β (Polβ). By varying reaction conditions, namely, Mg2+/Mn2+ replacement/combination and ionic strength decrease, we found that under certain conditions both Polβ and Polι can catalyze lesion bypass across both cis- and trans-BP adducts in the presence of physiological dNTP concentrations. Polβ and Polι catalyze gap filling trans-lesion synthesis in an error prone manner. By contrast, Polλ selectively introduced the correct dCTP opposite the modified dG in the case of cis-BP-dG adduct only, and did not bypass the stereoisomeric trans-adduct under any of the conditions examined. The results suggest that Polλ is a specialized polymerase that can process these kinds of lesions.
KW - Base excision repair
KW - Benzo[a]pyrene
KW - DNA polymerases
KW - Lesion bypass
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U2 - 10.1016/j.dnarep.2014.10.005
DO - 10.1016/j.dnarep.2014.10.005
M3 - Article
C2 - 25460917
AN - SCOPUS:84908377616
VL - 24
SP - 1
EP - 9
JO - DNA Repair
JF - DNA Repair
SN - 1568-7864
ER -