Differential H3K27ac ChIP-seq Analysis of HCT116 Cells In Response To Interferon Beta

To show the utility of our method, MEIRLOP, we performed a differential motif enrichment analysis of regulatory elements modulated by interferon beta (IFN-β) treatment in HCT116 cells as measured by H3K27ac ChIP-seq. Cell culture and treatment: HCT116 CMV-osTIR1 RAD21-mAC cells were obtained from Masato T. Kanemaki and cultured in McCoy’s 5A medium supplemented with 10% FBS. Cells were grown in a 37˚C incubator with 5% CO2. Cells were treated with 0.1% final DMSO for 6 hours and then treated with either IFN-β (1000 unit/ml) (n=2) for one hour or not further treated (n=2). ChIP-seq and Crosslinking: Crosslinking and ChIP-seq were performed as described in Heinz et al., 2018 with few adjustments. Briefly, cells were fixed directly by adding formaldehyde into media to a final concentration of 1% formaldehyde for 10min at room temperature and quenched with 125mM Glycine. Cells were then pelleted at 300g for 5min at 4˚C, washed twice with cold PBS (with 0.5% BSA), snap frozen in liquid nitrogen and stored at -80˚C. ChIP-seq was performed on 500,000 cells as described in Heinz et al., 2018. H3K27ac antibodies were obtained from Active Motif (cat#:39133). Libraries were single-end sequenced for 84bp to a depth of 5.5 - 8.6 reads on an Illumina NextSeq500 instrument. Differential ChIP-seq analysis: After sequencing and adapter trimming with FastP, 5.5M - 8.6M reads per library were aligned to the GRCh38 reference genome using bowtie2 at overall alignment rates of 98.5% - 99.5%. For each stimulation (n = 2) and control sample (n = 2), MACS2 was used to call peaks of median length 1kbp relative to a background input sample (of 16M reads). DiffBind was used to call differentially acetylated peaks between stimulated and unstimulated conditions.