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| Status |
Public on Jul 06, 2018 |
| Title |
Genome-wide analysis of 8-oxoguanine DNA glycosylase-1 after oxidative stress exposure |
| Organism |
Homo sapiens |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
We report the application of ChIP-Sequencing for profiling genome-wide distribution of 8-oxoguanine DNA glycosylase1 (OGG1, a DNA base excision repair protein), after oxidative stress exposure mediated by glucose oxidase (GOx) 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 associated with peaks to 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 guanines (primarily 8-oxoG). OGG1 increased significantly at regulatory regions of CXC-, CC- chemokines, cytokines and growth factors. In controls, the DNA was chromatin immunoprecipitated using antibody to NF-κB/RelA. As expected NF-κB was enriched on gene regulatory regions including those of proinflammatory,cell proliferation ones. These and other data derived from further analysis, suggest that OGG1 modulates gene expression in coordination with NF-κB.
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| Overall design |
FLAG-tagged OGG1 expressing HEK293 cells were stimulated with GOx (1 µU) for 0, 15, 30 and 60 min. After GOx exposure, protein-DNA complexes were crosslinked using formalin according to standard Millipore® protocol. DNA was sheared with 10-sec pulses using a Cole-Parmer GEX 130 ultrasonic processor (average 300 bp) and chromatin was immunoprecipitated (IP) with antibody to FLAG or anti-RelA/p65 antibody and IPs were collected using G-agarose beads (Millipore Corporation). After extraction, the DNA was subjected to high-throughput sequencing using Solexa/Illumina genome analyzer (Ambry Genetics).
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| Contributor(s) |
Aguilera-Aguirre L, Boldogh I |
| Citation(s) |
29940424 |
| Submission date |
Oct 21, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Istvan Boldogh |
| Organization name |
UNIVERSITY OF TEXAS MEDICAL BRANCH
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| Department |
Microbiology & Immunology
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| Street address |
301 UNIVERSITY BLVD.
|
| City |
Galveston |
| State/province |
Texas |
| ZIP/Postal code |
77555 |
| Country |
USA |
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| Platforms (1) |
| GPL15433 |
Illumina HiSeq 1000 (Homo sapiens) |
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| Samples (8)
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| Relations |
| BioProject |
PRJNA349805 |
| SRA |
SRP091905 |
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