Mutagenicity and Sequence Specificity of Acrolein-DNA Adduct

摘要

Acrolein (Acr) is a major toxicant in cigarette smoke (CS); it can interact with DNA forming two major adduct isomers: α-OH-Acr-dG and γ-OH-Acr-dG. Previously, we found that the Acr-DNA binding pattern in the human p53 gene coincides with the p53 mutational pattern in CS-related lung cancer; hence we proposed that Acr is a major lung cancer etiological agent (). This hypothesis has been brought into question with recent work that failed to detect Acr-induced mutations in the pSP189 system (). To resolve this controversy, we determined the level and the type of Acr-dG formation, and the mutagenicity of Acr-dG adducts in the same pSP189 system. We also mapped the Acr-dG adduct distribution at the nucleotide level, and the Acr-dG induced mutational spectrum in this system. We found that 1) γ-OH-Acr-dG is the major adduct formed in Acr-modified DNA based on the LC-ESI-MS/MS analysis; 2) the mutation frequency is proportional to the extent of Acr-modifications and the majority of which are G:C to T:A and G:C to A:T mutations; and 3) sequences with a run of G’s are the mutational hotspots. Using the UvrABC nuclease incision method to map the Acr-dG distribution in the supF gene sequence, we confirmed that Acr-DNA adducts preferentially form in guanine-rich sequences that are also mutational hotspots. These results reaffirm that Acr-dG adducts are mutagenic, and support our hypothesis that Acr is a major etiological agent for CS and cooking fume-related lung cancer.