Genome-Wide Mapping of Oxidative DNA Damage via Engineering of 8-Oxoguanine DNA Glycosylase

Abstract

The occurrence of 8-oxo-7,8-dihydroguanine (OG) in the genome, as one of the major DNA oxidative damages, has been implicated in an array of biological processes, ranging from mutagenesis to transcriptional regulation. Genome-wide mapping of oxidative damages could shed light on the underlying cellular mechanism. In the present study, we engineered the hOGG1 enzyme, a primary 8-oxoguanine DNA glycosylase, into a guanine oxidation-profiling tool. Our method, called enTRAP-seq, successfully identified more than 1400 guanine oxidation sites in the mouse embryonic fibroblast genome. These OG peaks were enriched in open chromatin regions and regulatory elements, including promoters, 5′ untranslated regions, and CpG islands. Collectively, we present a simple and generalizable approach for the genome-wide profiling of DNA damages with high sensitivity and specificity.

Genome-wide analysis of 8-oxo-7,8-dihydro-2'-deoxyguanosine at single-nucleotide resolution unveils reduced occurrence of oxidative damage at G-quadruplex sites

“…In this respect, CLAPS-seq and OGG1-AP-seq ( 24 ) performed damage recognition immediately after DNA extraction by directly labeling with biotin, or converting OGs to AP sites which were then biotinylated with ARP, respectively, thus these methods should suffer from less incidental oxidation. In contrast, OG-seq ( 22 ), click-code-seq ( 25 ), enTRAP-seq ( 23 ) and OxiDIP-seq ( 21 ) all sheared DNA before damage recognition/labeling. OxiDIP-seq had an extra heat-denaturing step (5 min at 95 °C) before immunoprecipitation ( 21 ).…”

“…In this regard, a couple of high-throughput sequencing-based techniques measuring the genomic distribution of OG in yeast, mouse and human genomes were developed recently ( 21–25 ). These methods rely on the recognition of OG by an anti-OG antibody ( 21 ), selective chemical reaction ( 22 ), or OG-specific glycosylases ( 23–25 ). Each method has its own advantages and disadvantages ( 26 ).…”

Next-generation DNA damage sequencing

“…As another example, an OGG1 K249Q mutant lacking glycosylase activity was used to trap a stable complex of OGG1 with the sequences containing 8-oxodG (enTRAP-seq). 57 Following affinity precipitation and sequencing, enTRAP-seq revealed enrichment of 8-oxodG in transcriptionally active chromatin regions and regulatory elements such as promoters, 5 0 UTRs, and CpG islands in the mouse embryonic fibroblast genome. While 8-oxodG-specific binding proteins are useful tools for 8-oxodG enrichment and sequencing, further studies comparing the binding specificity of antibody clones and glycosylase mutants will help to understand apparently conflicting results.…”

Genome-wide mapping of genomic DNA damage: methods and implications

“… Enzyme-mediated trapping and affinity precipitation of damaged DNA and sequencing (enTRAP-Seq) is the latest method developed for mapping 8-oxodG at the genome-wide level. This method was published by Zou’s group [ 69 ] and was used to map 8-oxodG across the mouse genome (i.e. in MEFs).…”