Fig. 2: Genome-wide multiomics analysis of OCPs in genotoxic stress-influenced cell models. | Nature Communications

Fig. 2: Genome-wide multiomics analysis of OCPs in genotoxic stress-influenced cell models.

From: Proteomic profiling and genome-wide mapping of O-GlcNAc chromatin-associated proteins reveal an O-GlcNAc-regulated genotoxic stress response

Fig. 2

a Immunoblotting and silver staining of GalNAz labeled MCF-7 and ADR cell lysates and chromatin. Chromatin proteins of the cells in the presence of 1 mM GalNAz (24 h) were extracted and reacted with alkyne-biotin. The cellular O-GlcNAc and expression of OGT were analyzed by immunoblotting. All blots and sliver staining are representative of at least two biologically independent experiments. Source Data are provided as a Source Data file. b Scatter plot showing the RNA-seq DEG expression levels (fragments per kilobase of transcript per million mapped reads (FPKM), fold change ≥ 2, FDR ≤ 0.001) in MCF-7 vs. ADR cells. Genes mentioned in this study are labeled. n = 2 biologically independent RNA-seq replicates. c GSEA of RNA-seq DEGs in MCF-7 cells compared with ADR cells. The normalized enrichment score (NES) and p value are indicated. d Schematic of the multiomics strategy involving a genome-wide chemical reporter-based method. MCF-7 cells were exposed to stepwise increasing concentrations of Adm for 8 months to generate the genotoxic resistant variant ADR cells. MCF-7 and ADR cells were incubated on 1 mM GalNAz media for 24 h. Decrosslinked OCPs were compared by label-free relative quantitative proteomics. The genomic DNA fragments bounded by OCPs were decrosslinked and then subjected to next-generation sequencing (COGC-seq). DEGs between MCF-7 and ADR cells were identified by RNA-seq analysis. An overlap analysis of three layered omics datasets was subsequently performed to uncover the O-GlcNAc regulated genotoxic stress-responsive reprograming. e Volcano plot of label-free relative quantitative proteomics data of OCPs in MCF-7 and ADR cells (n = 9 biologically independent experiments, two-sided unpaired Student’s t-test). The OCPs mentioned in this study are labeled. f Coverage plot of the COGC-seq peak locations over the whole human genome. The peaks of ADR cells showed distinct characteristic with those of MCF-7 cells. g Heat maps of the COGC-seq signal density at the peak center and TSSs (±3 kb). The average signal profile is shown. The red color indicates low a signal, and a blue color indicates a high signal.

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