DNA oxidation by reactive oxygen species typically occurs on guanine (G) to yield the base modification 8-oxo-7,8-dihydroguanine (OG), and G-rich, G-quadruplex (G4) sequences are particularly prone to oxidation.  Previously we identified reactive sites on the VEGF promoter G4 sequence toward oxidation to yield OG as a product.  The presence of OG at core sites can disrupt the G4 fold when the main four G runs of this sequence are studied while the addition of the native fifth G run (i.e., spare tire) allows maintenance of the folded state in the presence of disruptive OG.  From this observation, we hypothesized oxidation of G to OG could induce the VEGF promoter G4 to switch structures and impact gene expression.  Herein, the VEGF promoter G4 with four or five G runs containing a single OG at one of the reactive sites was placed in the promoter of a luciferase reporter gene to monitor transcription in mouse cells.  Luciferase expression for the system with four guanine runs was impacted by the presence of OG compared to the native VEGF sequence. Movement of the OG to other reactive sites found the extent of gene expression to depend on how disruptive the OG was to the four G run VEGF G4 sequence based on thermal melting studies.  When the fifth guanine track was added, gene expression increased when OG was present and the positional dependency in OG on expression was minimized.  This observation supports our hypothesis that the fifth G run can operate in the mode of a spare tire to maintain G4 folding when disruptive modifications to Gs are present.  Comparison of this system to other gene regulatory base modifications is discussed.

DNA base modification 8-oxo-7,8-dihydroguanine in a gene promoter regulates transcription

252nd National Meeting (2016)

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