Human DNA polymerase λ catalyzes lesion bypass across benzo[a]pyrene-derived DNA adduct during base excision repair

• Published in: DNA repair 2012-04, Vol.11 (4), p.367-373
• Main Authors: Skosareva, Lidia V., Lebedeva, Natalia A., Rechkunova, Nadejda I., Kolbanovskiy, Alexander, Geacintov, Nicholas E., Lavrik, Olga I.
• Format: Article
• Language: English
• Subjects: AP site processing; Base excision repair; Benzo(a)pyrene - chemistry; Benzo(a)pyrene - metabolism; Biocatalysis; DNA Adducts - genetics; DNA Adducts - metabolism; DNA Polymerase beta - metabolism; DNA polymerase λ; DNA Primers - genetics; DNA Primers - metabolism; DNA Repair; DNA-(Apurinic or Apyrimidinic Site) Lyase - metabolism; Humans; Hydrolysis; Isomerism; Lesion bypass
• ISSN: 1568-7864; 1568-7856
• DOI: 10.1016/j.dnarep.2012.01.002

► We examine the repair of an AP site in the context of clusters with bulky lesions. ► AP site opposite the cis-isomeric BP-dG-adduct is repaired using Polλ. ► AP site in adjacent 3′ (+1) position is the most difficult to repair. ► In position +1, dCTP is preferably inserted substrate for Polλ. 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 residues in certain positions were annealed with complementary oligonucleotides bearing either a cis- or trans-BP adduct. The resulting DNA duplexes contained dU either directly opposite the modified dG or shifted to adjacent 5′ (−1) or 3′ (+1) positions. Digestion with uracil DNA glycosylase was utilized to generate AP sites which were then hydrolyzed by APE1, and the resulting gaps were processed by DNA polymerase β (Polβ) or λ (Polλ). The AP sites in position −1 can be repaired effectively using APE1 and Polβ or Polλ. The AP sites opposite the BP lesions can be repaired using Polλ in the case of cis- but not the trans-isomeric adduct. The AP sites in position +1 are the most difficult to repair. In the case of the AP site located in position +1, the activity of Polλ does not depend on the stereoisomeric properties of the BP lesions and dCTP is the preferred inserted substrate in both cases. The capability of Polλ to introduce the correct dNTP opposite the cis-BP-dG adduct in gap filling reactions suggests that this polymerase may play a specialized role in the process of repair of these kinds of lesions.