8-Oxo-7,8-dihydroguanine in the Context of a Gene Promoter G-Quadruplex Is an On-Off Switch for Transcription

Abstract

Interplay between DNA repair of the oxidatively modified base 8-oxo-7,8-dihydroguanine (OG) and transcriptional activation has been documented in mammalian genes. Previously, we synthesized OG into the VEGF potential G-quadruplex sequence (PQS) in the coding strand of a luciferase promoter to identify that base excision repair (BER) unmasked the G-quadruplex (G4) fold for gene activation. In the present work, OG was site-specifically synthesized into a luciferase reporter plasmid to follow the time-dependent expression in mammalian cells when OG in the VEGF PQS context was located in the coding vs template strands of the luciferase promoter. Removal of OG from the coding strand by OG glycosylase-1 (OGG1)-mediated BER upregulated transcription. When OG was in the template strand in the VEGF PQS context, transcription was downregulated by a BER-independent process. The time course changes in transcription show that repair in the template strand was more efficient than repair in the coding strand. Promoters were synthesized with an OG:A base pair that requires repair on both strands to yield a canonical G:C base pair. By monitoring the up/down luciferase expression, we followed the timing of repair of an OG:A base pair occurring on both strands in mammalian cells in which one lesion resides in a G-quadruplex loop and one in a potential i-motif. Depending on the strand in which OG resides, coding vs template, this modification is an up/downregulator of transcription that couples DNA repair with transcriptional regulation.