Effects of the stimuli-dependent enrichment of 8-oxoguanine DNA glycosylase1 on chromatinized DNA

Abstract

8-Oxoguanine DNA glycosylase 1 (OGG1) initiates the base excision repair pathway by removing one of the most abundant DNA lesions, 8-oxo-7,8-dihydroguanine (8-oxoG). Recent data showed that 8-oxoG not only is a pro-mutagenic genomic base lesion, but also functions as an epigenetic mark and that consequently OGG1 acquire distinct roles in modulation of gene expression. In support, lack of functional OGG1 in Ogg1-/- mice led to an altered expression of genes including those responsible for the aberrant innate and adaptive immune responses and susceptibility to metabolic disorders. Therefore, the present study examined stimulus-driven OGG1-DNA interactions at whole genome level using chromatin immunoprecipitation (ChIP)-coupled sequencing, and the roles of OGG1 enriched on the genome were validated by molecular and system-level approaches. Results showed that signaling levels of cellular ROS generated by TNFα, induced enrichment of OGG1 at specific sites of chromatinized DNA, primarily in the regulatory regions of genes. OGG1-ChIP-ed genes are associated with important cellular and biological processes and OGG1 enrichment was limited to a time scale required for immediate cellular responses. Prevention of OGG1-DNA interactions by siRNA depletion led to modulation of NF-κB's DNA occupancy and differential expression of genes. Taken together these data show TNFα-ROS-driven enrichment of OGG1 at gene regulatory regions in the chromatinized DNA, which is a prerequisite to modulation of gene expression for prompt cellular responses to oxidant stress.

Enzymatically inactive OGG1 binds to DNA and steers base excision repair toward gene transcription

“…HEK 293 cells were over‐expressed with FLAG‐tagged wt OGG1 and the mutants, and then, exposed to TNFα for 60 minutes. ChIP was performed and the enrichment of OGG1 at promoter sequences was determined using quantitative PCR (qPCR) as described previously 33,34 . The right panel of Figure 4A shows that wt OGG1 and the mutants were expressed equally in transfected HEK 293 cells.…”

“…OGG1 oxidative modification at cysteine residues is a reversible modification that can be regained after redox balance is restored 25,33,36,37 . Inactivation of OGG1 via cysteine oxidation does not interfere with genomic substrate binding, but halts its glycosylase activity 3,23,25,33‐34,39 . The vast numbers of OGG1 K249Q analyses help us to comprehend the mechanism of how oxidatively inactivated OGG1 modulates gene expression.…”

“…Cysteine‐oxidized OGG1 persists its ability to recognize the substrate, bend DNA and extrude the damaged base into the active pocket; however, the conformation of the enzyme/DNA complex does not accommodate for base excision, probably due to the disfavor of the lysine 249 to attack C1′ of the deoxyribose moiety using its ε‐amino group 51 . The biological significance of OGG1 oxidation was proclaimed by studies suggesting that the pre‐excision steps and the associated adjustment of the adjacent DNA by oxidized OGG1 facilitate DNA occupancy of TFD II, Sp1, and NF‐κB, and consequently enhance expression of pro‐inflammatory genes ( TNF , CXCLs , and ILs ) under OS conditions 3,25,33‐34,39,48 . Upon normalization of the cellular redox state, genome fidelity was accomplished via DNA‐BER 33,36,37 .…”

“…The size of the PCR product is ~200 bp. The fold enrichment of wtOGG1 or its mutants on the specific region of genomic DNA was calculated as described previously 33,34 …”

Protector properties of N-acetylcysteine, melatonin and their compatible use for oxidative damage of rat brain cells with experimental diabetes mellitus type 1

“…Визначення вмісту 8-oxoG в біологічних середовищах є високоінформативним для оцінки рівня окисного пошкодження ДНК [24]. Повідомляється про зростання рівня 8-oxoG у тканинах головного мозку при нейродегенеративних захворюваннях [25,26].…”