Induction of action-at-a-distance mutagenesis by 8-oxo-7,8-dihydroguanine in DNA pol λ-knockdown cells

INTRODUCTION: In DNA, 8-oxo-7,8-dihydroguanine (G(O), 8-hydroxyguanine) is one of the most pivotal oxidatively damaged bases and induces G:C → T:A transversion mutations. DNA polymerase λ preferentially inserts dCTP opposite G(O)in vitro, and this error-free bypass function is considered to be important after A base removal from G(O):A pairs by the MUTYH DNA glycosylase. To examine the effects of reduced levels of DNA polymerase λ on the G(O)-induced mutations, the polymerase was knocked-down in human U2OS cells, and a shuttle plasmid DNA containing a G(O):C pair at position 122 in the supF gene was transfected into the cells. The plasmid DNA replicated in the cells was introduced into an Escherichia coli indicator strain, to measure the supF mutant frequency. RESULTS: The knockdown of DNA polymerase λ significantly enhanced the mutant frequency of the G(O) plasmid DNA. Contrary to our expectations, the knockdown did not promote the targeted G:C → T:A transversion. Instead, substitution mutations at G:C sites other than position 122 were enhanced in the cells. CONCLUSIONS: These results suggested that the knockdown of DNA polymerase λ induced action-at-a-distance mutagenesis in human cells when the G(O):C pair was present in the DNA.