P12 - 8-OxoG Formation: Impact in Gene Promoters on Transcription and Mapping Studies

Abstract

Studies with the regulatory VEGF promoter G-quadruplex (G4)-forming sequence have identified this G-rich region is prone to oxidation yielding 8-oxoguanine (OG),1 and we postulated that oxidation of the regulatory sequence could alter gene expression. We used synthetic chemistry to install the VEGF G4 with OG at oxidation-prone sites in the promoter of a luciferase gene that was transfected in mammalian cells to understand how gene expression changed. The modified sequences were installed in either the native coding strand or the non-native template strand of the promoter.1,2 Monitoring the time dependency in gene expression found a nearly 300% increase in expression when OG was in the coding strand and a nearly 400% decrease in expression for OG in the template strand. Studies in base excision repair (BER) proficient or deficient cell lines support BER as the process for gene activation for OG in the coding strand, and template strand OG turned gene expression off in a BER-independent process. Furthermore, we found that human DNA repair gene promoters are favorably biased with potential G4-forming sequences that can alter gene expression when OG is present. Thus, we hypothesize that certain potential G4-forming sequences are switches for gene expression during oxidative stress.3 To determine whether OG is prevalent in gene regulatory regions we developed a sequencing protocol and found enrichment of OG in gene promoters and 5'-UTRs.4