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Evidence of complete cellular repair of 1,N6-ethenoadenine, a mutagenic and potential damage for human cancer, revealed by a novel method

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Abstract

1,N6-Ethenoadenine (εA) is generated endogenously by lipid peroxidation and exogenously by tumorigenic industrial agents, vinyl chloride, and vinyl carbamate. εA detected in human tissues causes mutation and is implicated in liver, colon and lung cancers. N-methyl purine DNA-glycosylase (MPG) is the only enzyme known so far to repair εA. However, the mechanism of in vivo repair of εA and the role of MPG remain enigmatic. Moreover, previous in vivo repair studies for DNA lesions, including εA, focused only on the step of the removal of the base lesion without further insight into the completion of the repair process. This may be in part due to the unavailability of an appropriate in vivo quantitative method to evaluate complete BER process at the basal level. Our newly developed in vivo method is highly sensitive and involves phagemid M13mp18, containing εA at a defined position. The complete repair events have been estimated by plaque assay in E. coli with the phagemids recovered from the human cells after cellular processing. We found that the detectable complete (removal and replacement of εA with adenine) repair was observed only 18% in 16 h, but with the repair nearing completion within 24 h in colon cancer, HCT-116, cells. Moreover, MPG is the predominant enzyme for the BER process to remove εA in mammalian cells. Although, the εA is fairly a bulky adduct compared to other small BER substrate lesions, NER pathway is not involved in repair of this adduct. Furthermore, the εA repair in vivo and in vitro is predominant in the G0/G1 phase of the cell cycle.

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Abbreviations

BER:

Base excision repair

MPG:

N-Methylpurine DNA glycosylase

Hx:

Hypoxanthine

εA:

1,N6-Ethenoadenine

m3A:

3-Methyl adenine

m7G:

7-Methyl guanine

CCC:

Covalently closed circular

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Acknowledgments

We thank Dr. Leona Samson (MIT) for providing the MPG−/− and +/+ MEFs. We thank Ms. Karen Howenstein for expert editorial and administrative help. The work was supported by NIH grant RO1 CA 92306 (RR).

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Authors and Affiliations

  1. Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, Washington, DC, 20057, USA

    Sujata Choudhury, Sanjay Adhikari, Amrita Cheema & Rabindra Roy

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  1. Sujata Choudhury
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  2. Sanjay Adhikari
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  4. Rabindra Roy
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Correspondence to Rabindra Roy.

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Choudhury, S., Adhikari, S., Cheema, A. et al. Evidence of complete cellular repair of 1,N6-ethenoadenine, a mutagenic and potential damage for human cancer, revealed by a novel method. Mol Cell Biochem 313, 19–28 (2008). https://doi.org/10.1007/s11010-008-9737-1

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