The Mechanism Of DNA Repair Enzyme OGG1 Regulates Inflammatory Genes Expression

Biological aerobic respiratory metabolism and environmental factors can lead to the increase of intracellular reactive oxygen species(ROS).The production of ROS in cells is higher than the clearance rate of antioxidant enzymes,which is called oxidative stress.Oxidative stress causes oxidation of macromolecule substances.When proteins,lipids and RNA suffered oxidatived modification,they will be degraded or reused by cells.DNA damage must be repaired in order to ensure the integrity and fidelity of the genome.Because the redox potential of guanine(G)is the lowest,the main oxidative damage product of DNA is 8-hydroxyguanine(8-oxoG).In eukaryotic cells,8-hydroxyguanine DNA glycosylase(OGG1)is the main DNA repair protein that specifically exicision 8-oxoG on DNA.OGG1 recognizes and removes 8-oxoG through the initial base excision repair pathway(OGG1-BER),and then recruits other proteins to complete the gap.During DNA replication,8-oxoG can be transpaired with cytosine(C)or cis-paired with adenine(A),so it is possible to replace GC-paired into TA-paired mutations after two cell cycles.Therefore,the accumulation of 8-oxoG in the genome or the abnormal repair ability of OGG1 are usually considered as the main inducement of cell aging or canceration.Although guanine is easy oxidized and 8-oxoG is a potential mutagenic site,vertebrate genes tend to be distributed in regions with high GC content,and more than 72% of human gene promoters have high GC content.In recent years,more and more evidences showed that 8-oxoG and its specific repair protein OGG1 play epigenetic roles in gene transcription.Our previously work showed that when ROS attacked cells,oxidized OGG1 binding to DNA induced DNA bends,facilitates the transcription factor NF-?B binding to gene promoter regions to promotes the transcription of inflammatory factors CXCL1 and TNF?.However,whether OGG1 promotes gene transcription depends on its digestion activity,the distribution of OGG1 at the whole genome level,OGG1 involved biological process and whether OGG1 can be used as a target to prevent and treat diseases caused by oxidative stress remain to be further revealed.This study: 1)utilized OGG1 and its mutant plasmids to do EMSA,qRT-PCR,RNA FISH,ChIP,Luciferase assay and other experiments.Results showed that the OGG1 K249 Q mutation(only have binding activity without cleavage activity)could bind and activate Cxcl2 promoter to promote gene transcription,while OGG1 C253A(weak binding and cleavage activity)had no significant effect on the activation of Cxcl2 promoter and Cxcl2 gene expression upon oxidative response.These data revealed that OGG1 promotes the expression of inflammation genes independent of its cleavage activity;2)By ChIP-Sequencing,IGV and Gorilla software analysis and combined with qRT-PCR,ChIP-qPCR and EMSA techniques,we found that OGG1 mainly binds to the gene regulatory region at whole genome level and participates in the regulation of inflammation(P=5.51E-07),oxidative stress(P=1.04E-23),and the negative regulation of programmed cell death(P=1.97E-11),positive regulation of cell metabolism(P=4.15E-09)and other important biological processes;3)Designing a small molecule compound TH5487 according to the active site of OGG1.By utilized qRT-PCR,ChIP,EMSA,mouse nose dropping,Giemsa staining and HE staining experiments,we found that TH5487 can prevent OGG1 binding to DNA,thereby decreasing NF-?B binding to inflammatory factors promoters(have no affect of NF-?B binding to DNA)and effectively inhibits the recruitment of neutrophils to the mice lungs,thus inhibits the occurrence of acute pulmonary inflammation.O8 is another inhibitor of OGG1(have no affect of OGG1-DNA binding,but decrease its cleavage activity)and have no affect of inflammatory factors expression.This research further reveals the molecular mechanism of OGG1 as an epigenetic regulator of gene expression via a series of basic research technology.And the small molecule compounds designed by targeting OGG1 can treat the inflammatory response caused by oxidative stress,providing a good prospect for the treatment of other diseases caused by OGG1(oxidative stress).