Carcinogens often generate mutable DNA lesions that contribute to cancer and aging. However, the chemical structure of tumorigenic DNA lesions formed by acetaldehyde remains unknown, although it has long been considered an environmental mutagen in alcohol, tobacco, and food. Here, we identify an aldehyde-induced DNA lesion, forming an intrastrand crosslink between adjacent guanine bases, but not in single guanine bases or in other combinations of nucleotides. The GG intrastrand crosslink exists in equilibrium in the presence of aldehyde, and therefore it has not been detected or analyzed in the previous investigations. The newly identified GG intrastrand crosslinks might explain the toxicity and mutagenicity of acetaldehyde in DNA metabolism.

Background: Acetaldehyde, produced upon exposure to alcohol, cigarette smoke, polluted air and sugar, is a highly reactive compound that is carcinogenic to humans and causes a variety of DNA lesions in living human cells. Previously, we reported that acetaldehyde reacts with adjacent deoxyguanosine residues on oligonucleotides, but not with single deoxyguanosine residues or other deoxyadenosine, deoxycytosine, or thymidine residues, and revealed that it forms reversible intrastrand crosslinks with the dGpdG sequence (GG dimer). Results: Here, we show that restriction enzymes that recognize a GG sequence digested acetaldehyde-treated plasmid DNA with low but significant efficiencies, whereas restriction enzymes that recognize other sequences were able to digest such DNA. This suggested that acetaldehyde produced GG dimers in plasmid DNA. Additionally, acetaldehyde-treated oligonucleotides were efficient in preventing digestion by the exonuclease function of T4 DNA polymerase compared to non-treated oligonucleotides, suggesting structural distortions of DNA caused by acetaldehyde-treatment. Neither in vitro DNA synthesis reactions of phi29 DNA polymerase nor in vitro RNA synthesis reactions of T7 RNA polymerase were observed when acetaldehyde-treated plasmid DNA was used, compared to when non-treated plasmid DNA was used, suggesting that acetaldehyde-induced DNA lesions inhibited replication and transcription in DNA metabolism. Conclusions: Acetaldehyde-induced DNA lesions could affect the relative resistance to endo-and exo-nucleolytic activity and also inhibit in vitro replication and in vitro transcription. Thus, investigating the effects of acetaldehydeinduced DNA lesions may enable a better understanding of the toxicity and carcinogenicity of acetaldehyde.

Adenine bases in DNA, RNA, and nucleotides are deaminated during normal metabolism via hydrolytic and nitrosative reactions. In RNA, the deaminated product inosine is resolved by human endonuclease V, and mice deficient in this enzyme are cancer-prone. We have now produced, purified, and characterized naturally occurring variants of human endonuclease V (V29I, R112Q, K114R, H141Y, and D201N). We found that H141Y, but not other variants, is catalytically impaired, suggesting that individuals homozygous for H141Y may be predisposed to disease.