Project description:7,8-dihydro-8-oxoguanine (OG) is one of the major DNA oxidative damages, which has been concerned by researchers for its potential of mutagenesis and epigenetic regulation. Genome-wide mapping of oxidative DNA damage is of great value for understanding the biological impact of these modifications. Here we present a novel appoach to profile guanine oxidation in the mouse genome. Through structure-guided mutagenesis, we've successufully engineered 8-oxoguanine DNA glycosylase into a useful tool for OG detection. By enrichment of glycosylase-DNA complex, thousands of OG peaks were identified in the mouse genome.
Project description:8-oxoguanine is excised from DNA by 8-oxoguanine DNA glycosylase-1 during DNA base excision repair. We used microarrays to gain insight into the potential biological function of the excised, free 8-oxoguanine base. MRC5, human diploid fibroblasts were exposed to 8-oxoguanine and changes in gene expression compared to mock control were analyzed 6 hours after exposure. MRC5, human diploid fibroblasts were exposed to 8-oxoguanine and changes in gene expression compared to mock control were analyzed 12 hours after exposure.
Project description:Balb/cJ mouse received control RNAi or RNAi to 8-oxoguanine DNA glycosylase (Ogg1) intranasally prior to the exposure for 1 h to a) glucose oxidase (1 mU) to induce OGG1-BER or b) OGG1-BER product 8-oxoguanine (1 μM). We used SABiosciences Mouse Inflammatory Cytokines & Receptors PCR Array (PAMM-011A) to quantitate inflammatory gene expression dependent on OGG1 and its product 8-oxoguanine.
Project description:8-oxoguanine is excised from DNA by 8-oxoguanine DNA glycosylase-1 during DNA base excision repair. We used microarrays to gain insight into the potential biological function of the excised, free 8-oxoguanine base.
Project description:Altered oncogene expression in cancer cells causes loss of redox homeostasis resulting in oxidative DNA damage, e.g., 8-oxoguanine (8-oxoG), repaired by base excision repair (BER). PARP1 coordinates BER and relies on the upstream 8-oxoguanine-DNA glycosylase (OGG1) to recognise and excise 8-oxoG. Here we hypothesize that OGG1 may represent an attractive target to exploit reactive oxygen species (ROS) elevation in cancer. Although OGG1 depletion is well tolerated in non-transformed cells, we report here that OGG1 depletion obstructs A3 T-cell lymphoblastic acute leukemia growth in vitro and in vivo, validating OGG1 as a potential anti- cancer target. In line with this hypothesis, we show that OGG1 inhibitors (OGG1i) target a wide range of cancer cells, with a favourable therapeutic index compared to non-transformed cells. Mechanistically, OGG1i and shRNA depletion cause lost mitochondrial function leading to S- phase DNA damage, replication stress and proliferation arrest or cell death, representing a novel mechanistic approach to target cancer. This study adds OGG1 to the list of BER factors, e.g., PARP1, as potential targets for cancer treatment. _x000B_
Project description:Balb/cJ mouse received control RNAi or RNAi to 8-oxoguanine DNA glycosylase (Ogg1) intranasally prior to the exposure for 1 h to a) glucose oxidase (1 mU) to induce OGG1-BER or b) OGG1-BER product 8-oxoguanine (1 M-NM-<M). We used SABiosciences Mouse Inflammatory Cytokines & Receptors PCR Array (PAMM-011A) to quantitate inflammatory gene expression dependent on OGG1 and its product 8-oxoguanine. After intranasal challenge with glucose oxidase or 8-oxoguanine, mice were sacrificed and lungs were collected and processed to obtain RNA. Total pooled (n=5) RNA (1 M-NM-<g) was reverse transcribed into cDNA using SuperscriptM-BM-. III First Strand Synthesis System (Invitrogen), mixed with equal amounts of 2X SYBR Green Supermix (Qiagen) and 20 M-NM-<l of reaction mixture was added to each well of the PAM-011A array. The reaction was evaluated using an ABI PRISMM-BM-. 7000 Sequence Detection System using recommended settings by SABiosciences.
Project description:We report the application of ChIP-Sequencing for profiling genome-wide distribution of 8-oxoguanine DNA glycosylase (OGG1) which is a DNA base excision repair protein, after TNF exposure of cells. By obtaining an average of over 18 million of total reads per sample and over 19.5 million of unique reads per sample from chromatin immunoprecipitated DNA, we generated genome-wide chromatin-state maps of HEK293 cells. We performed gene ontology and functional pathway analysis of genes associate with peaks, and identify regions of the genomes (e.g. promoter, introns, etc) where the peaks tend to occur. The results show that OGG1 is primarily associated with promoter regions in vicinity of transcription factor binding sites 5' of transcription start sites (TSS). This pattern of distribution occurs in spite of genome-wide oxidative modifications of guanine (primarily 8-oxoG). OGG1 increased highly significant (1e-09 to 1e-517) enrichment of 57 transcription factor binding sites including those for Sox(2, 3, 6, 10), RUNX, RUNX(1, 2), HOX(C, D), STAT(1, 3, 6), IRF(1 to 4), NF-κB. In controls, the DNA was chromatin immunoprecipitated using antibody to NF-κB/RelA. These and other data derived from further analysis, suggest that OGG1 modulates binding of transcription factors and gene expression.
Project description:To assess the role of 8-oxoguanine DNA glycosylase 1 (OGG1) on TNF induced pro-inflammatory gene expression. We used Qiagen Human Inflammatory Cytokines & Receptors PCR Array (PAHS-011Z) to quantitate inflammatory gene expression dependent on OGG1.