Mutagenesis by acrolein-derived propanodeoxyguanosine adducts in human cells

Acrolein, which is widely spread in the environment and is produced by lipid peroxidation in cells, reacts with DNA to form two exocyclic 1,N²-propanodeoxyguanosine (PdG) adducts. To establish their relative contribution to the acrolein mutagenicity, the genotoxic properties of α-OH-PdG and γ-OHP-dG together with their model DNA adduct, PdG, were studied in human cells. DNA adducts were incorporated site-specifically into a SV40/BK virus origin-based shuttle vector and replicated in xeroderma pigmentosum complementation group A (XPA) cells. Analysis of progeny plasmid revealed that α-OH-PdG and PdG strongly block DNA synthesis and that both adducts induced base substitutions with G → T transversions predominating. Primer extension studies, catalyzed by the 3′→5′ exonuclease-deficient Klenow fragment of Escherichia coli pol I, revealed limited extension from the 3′ primer termini opposite these two adducts. In contrast, γ-OH-PdG did not strongly block DNA synthesis or miscode in XPA cells. Primer extension from a dC terminus opposite γ-OH-PdG was much more efficient than that opposite α-OH-PdG or PdG. These results indicate that the minor α-OH-PdG adduct is more genotoxic than the major γ-OH-PdG. Furthermore, experiments using a HeLa whole cell extract indicate that all three DNA adducts are not efficiently removed from DNA by base excision repair.