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3-Methyladenine DNA glycosylase is important for cellular resistance to psoralen interstrand cross-links.

ABSTRACT: DNA interstrand cross-links (ICLs), widely used in chemotherapy, are cytotoxic lesions because they block replication and transcription. Repair of ICLs involves proteins from different repair pathways however the precise mechanism is still not completely understood. Here, we report that the 3-methyladenine DNA glycosylase (Aag), an enzyme that initiates base excision repair at a variety of alkylated bases, is also involved in the repair of ICLs. Aag(-/-) mouse embryonic stem cells were shown to be more sensitive to the cross-linking agent 4,5',8-trimethylpsoralen than wild-type cells, but no more sensitive than wild-type to the psoralen derivative Angelicin that forms only monoadducts. We show that gamma-H2AX foci formation, a marker for double strand breaks that are formed during ICL repair, is impaired in psoralen treated Aag(-/-) cells in both quantity and kinetics. However, in our in vitro system, purified human AAG can neither bind to the ICL nor cleave it. Taken together, our results suggest that Aag is important for the resistance of mouse ES cells to psoralen-induced ICLs.

SUBMITTER: Maor-Shoshani A

PROVIDER: S-EPMC2583468 | biostudies-literature | 2008 Aug

REPOSITORIES: biostudies-literature

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3-Methyladenine DNA glycosylase is important for cellular resistance to psoralen interstrand cross-links.

Maor-Shoshani Ayelet A Meira Lisiane B LB Yang Xuemei X Samson Leona D LD

DNA repair 20080620 8

DNA interstrand cross-links (ICLs), widely used in chemotherapy, are cytotoxic lesions because they block replication and transcription. Repair of ICLs involves proteins from different repair pathways however the precise mechanism is still not completely understood. Here, we report that the 3-methyladenine DNA glycosylase (Aag), an enzyme that initiates base excision repair at a variety of alkylated bases, is also involved in the repair of ICLs. Aag(-/-) mouse embryonic stem cells were shown to ...[more]

PMID: 18571479

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Project description:Bifunctional DNA alkylating agents form a diverse assortment of covalent DNA interstrand cross-linked (ICL) structures that are potent cytotoxins. Because it is implausible that cells could possess distinct DNA repair systems for each individual ICL, it is believed that common structural and dynamic features of ICL damage are recognized, rather than specific structural characteristics of each cross-linking agent. Investigation of the structural and dynamic properties of ICLs that might be important for recognition has been complicated by heterogeneous incorporation of these lesions into DNA. To address this problem, we have synthesized and characterized several homogeneous ICL DNAs containing site-specific staggered N4-cytosine-ethyl-N4-cytosine cross-links. Staggered cross-links were introduced in two ways, in a manner that preserves the overall structure of B-form duplex DNA and in a manner that highly distorts the DNA structure, with the goal of understanding how structural and dynamic properties of diverse ICL duplexes might flag these sites for repair. Measurements of base pair opening dynamics in the B-form ICL duplex by (1)H NMR line width or imino proton solvent exchange showed that the guanine base opposite the cross-linked cytosine opened at least 1 order of magnitude more slowly than when in a control matched normal duplex. To a lesser degree, the B-form ICL also induced a decrease in base pair opening dynamics that extended from the site of the cross-link to adjacent base pairs. In contrast, the non-B-form ICL showed extensive conformational dynamics at the site of the cross-link, which extended over the entire DNA sequence. Because DNA duplexes containing the B-form and non-B-form ICL cross-links have both been shown to be incised when incubated in mammalian whole cell extracts, while a matched normal duplex is not, we conclude that intrinsic DNA dynamics is not a requirement for specific damage incision of these ICLs. Instead, we propose a general model in which destabilized ICL duplexes serve to energetically facilitate binding of DNA repair factors that must induce bubbles or other distortions in the duplex. However, the essential requirement for incision is an immobile Y-junction where the repair factors are stably bound at the site of the ICL, and the two DNA strands are unpaired.

Dataset Information

3-Methyladenine DNA glycosylase is important for cellular resistance to psoralen interstrand cross-links.

Publications

3-Methyladenine DNA glycosylase is important for cellular resistance to psoralen interstrand cross-links.

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