8-Oxo-7,8-dihydroguanine, friend and foe: Epigenetic-like regulator versus initiator of mutagenesis

Abstract

A high flux of reactive oxygen species during oxidative stress results in oxidative modification of cellular components including DNA. Oxidative DNA "damage" to the heterocyclic bases is considered deleterious because polymerases may incorrectly read the modifications causing mutations. A prominent member in this class is the oxidized guanine base 8-oxo-7,8-dihydroguanine (OG) that is moderately mutagenic effecting G→T transversion mutations. Recent reports have identified that formation of OG in G-rich regulatory elements in the promoters of the VEGF, TNFα, and SIRT1 genes can increase transcription via activation of the base excision repair (BER) pathway. Work in our laboratory with the G-rich sequence in the promoter of VEGF concluded that BER drives a shift in structure to a G-quadruplex conformation leading to gene activation in mammalian cells. More specifically, removal of OG from the duplex context by 8-oxoguanine glycosylase 1 (OGG1) produces an abasic site (AP) that destabilizes the duplex, shifting the equilibrium toward the G-quadruplex fold because of preferential extrusion of the AP into a loop. The AP is bound but inefficiently cleaved by apurinic/apyrimidinic endoDNase I (APE1) that likely allows recruitment of activating transcription factors for gene induction. The ability of OG to induce transcription ascribes a regulatory or epigenetic-like role for this oxidatively modified base. We compare OG to the 5-methylcytosine (5mC) epigenetic pathway including its oxidized derivatives, some of which poise genes for transcription while also being substrates for BER. The mutagenic potential of OG to induce only ∼one-third the number of mutations (G→T) compared to deamination of 5mC producing C→T mutations is described. These comparisons blur the line between friendly epigenetic base modifications and those that are foes, i.e. DNA "damage," causing genetic mutations.

Keywords: 8-Oxo-7,8-dihydroguanine; Base excision repair; Epigenetics; G-quadruplex; Mutagenesis; Oxidative stress.

Fig. 1

Comparison of the G oxidative modification cycle with the C methylation and oxidative modification cycle to illustrate the centrality of the abasic site (AP) to return the sequence back to the original active state. *For the sake of brevity, the 4-electron oxidation product of G, or 2-electron oxidation product of OG yielding 5-guanidinohydantoin (Gh) is shown; the other 4-electron product spiroiminodihydantoin (Sp) is not shown [7]. The yields of Gh and Sp show strong dependency on the reaction conditions and context, favoring Gh in duplex DNA oxidations or reactions at pH < 6 and favoring Sp in single-stranded and G-quadruplex DNA oxidations or reactions at pH > 7 [8, 9].

Fig. 2

Sequence for the PQS in the coding strand of the VEGF gene that upon oxidation of G to OG provides a substrate for BER that unmasks the G-quadruplex for gene induction. (A) The sequence of the G-rich element in the VEGF promoter. The Gs marked in red are sites in which OG was synthetically incorporated to demonstrate the proposed pathway in part D [14]. (B) Data illustrating the presence of OG in the VEGF promoter increased luciferase expression by >2.5 fold in MEF cells, and knocking out OGG1 results in the signal remaining unchanged relative to the wild type (WT) plasmid. (C) Utility of APE1-specific siRNAs in glioblastoma cells provided a dose response impact on luciferase expression. The data in panels B and C demonstrate OGG1 and APE1, respectively, are required for gene induction when OG is present in the VEGF promoter PQS. These data were adapted from the original publication [14]. (D) Proposed pathway for oxidation of the PQS to yield OG and guide the BER process by unmasking the G-quadruplex for gene activation, thus illustrating an intertwining of DNA repair and transcriptional induction.

Fig. 3

Structural comparison and base pairing properties of B-form and G4 DNA. The B-form DNA structure was derived from pdb 1BNA [40], and the VEGF G4 structure was derived from pdb 2M27 [31].

Fig. 4

Pathways to generate genomic mutations for OG and 5mC.