Fig 1.
Analysis of differentially expressed genes (DEGs) and differentially accessible regions (DARs) in mouse lung tissues.
(A) Overview of the study design. Four omics indicators with P. aeruginosa-infected lung tissue were integrated to dissect chromatin accessibility, transcripts, protein, and ubiquitinated protein abundance and alteration. (B) The average read density (top) and heatmap (bottom) representation of normalized ATAC-seq signals over DARs in the infection and control groups. Heatmaps display a read signal across 15,567 accessible regions. Signals within 1.5 kb of the center of DARs are shown in descending order. (C) Bar chart showing annotation of DARs to genomic features: 3′ UTR, 5′ UTR, exon, intergenic regions, introns, non-coding regions, promoter-TSS, and TTS. Promoter-TSS regions are defined as peak summits located up to 1 kb upstream and 100 bp downstream of the TSS. (D) Volcano plot showing the transcript levels of DEGs between the infection and control groups. Significantly upregulated 2126 genes are shown as red dots, whereas 2103 downregulated genes are shown as blue dots. Filtered by adjusted p value < 0.01, the top significantly up- and downregulated genes are highlighted in the text according to log2(FC). (E) Venn diagram showing the overlap between nearby genes from DARs and DEGs. (F) Correlation analysis between DARs and their nearest upregulated DEGs. Each blue dot represents a gene that is significantly differentially expressed and associated with chromatin accessibility changes. Based on log2(FC), the top-ranked DEGs are labeled and shown in the text. Pearson’s correlation coefficient (r) and the associated p value are displayed. (G), Key KEGG pathways associated with upregulated DEGs related to DARs.
Fig 2.
Identification and analysis of DEGs and DARs in P. aeruginosa-infected mouse alveolar macrophages.
(A) Overview of the study design. Two biological replicates infected with P. aeruginosa were evaluated to profile chromatin accessibility, and three biological replicates infected with P. aeruginosa were evaluated to profile transcript abundances. (B) The average read density (top) and heatmap (bottom) representation of the normalized ATAC-seq signal over DARs in the infection and control groups. Heatmaps show a read signal of 3403 available locations. Signals within 1.5 kb of the center of the DARs are shown in descending order. (C) Bar chart showing annotation of DARs to genomic features: 3′ UTR, 5′ UTR, exon, intergenic regions, introns, non-coding regions, promoter-TSS, and TTS. Promoter-TSS regions are defined as peak summits located up to 1 kb upstream and 100 bp downstream of the TSS. (D) Volcano plot showing the transcript levels of DEGs between the infection and control groups. A total of 2208 significantly upregulated genes are shown as red dots, whereas 1680 downregulated genes are shown as blue dots. Filtered by adjusted p value < 0.01, the top significantly up- and downregulated genes are highlighted in the text according to log2(FC). (E) Venn diagram showing the overlap between nearby genes from DARs and DEGs. (F) Heatmap depicting the expression levels of overlapping genes around upregulated DEGs and DARs.
Fig 3.
Molecular interactions between lung tissues and alveolar macrophages.
(A), Proportions of DARs within genomic regions in lung tissues and alveolar macrophages: 3′ UTR, 5′ UTR, exon, intergenic regions, introns, non-coding regions, promoter-TSS, and TTS. (B) Heatmap representation of normalized ATAC-seq signals of DARs shared by lung tissues and alveolar macrophages. (C) Correlation analysis of DEGs between lung tissues and mouse alveolar macrophage groups. Each blue dot represents a gene that is significantly differentially expressed and correlates with changes in chromatin accessibility. Pearson’s correlation coefficient (r) and associated p values are shown. (D) Pie chart showing the components of the ubiquitin-proteasome system of DEGs shared by mouse lung tissues and alveolar macrophages. (E) Bubble plot illustrating the significant enrichment of the ubiquitination function in the DEGs shared by mouse lung tissues and alveolar macrophages. The position corresponds to the number of genes, whereas color denotes significance. (F) IGV Browser view of four upregulated accessible chromatin regions of the components of the ubiquitin-proteasome system in alveolar macrophages and lung tissues.
Fig 4.
Identification and analysis of DEPs and DUPs in P. aeruginosa-infected lung tissues.
(A) The waterfall plot of the DEPs arrangement is sorted by log2(FC), with upregulated expression shown in red and downregulated expression shown in green. (B) The GO function enrichment analysis for upregulated DEPs is represented in a bubble plot. Color represents significance, and position represents the number of enriched genes. (C) The waterfall plot of the DUPs arrangement is sorted by log2(FC), with upregulated expression shown in yellow and downregulated expression shown in blue. (D) Results of the study of GO functions that are enriched for upregulated DUPs are displayed in a bubble plot. Position indicates the number of enriched genes, whereas color denotes significance. (E) Heatmap showing the abundance of molecules with altered transcriptomics, proteomics, and ubiquitin proteomics in the positive regulation of cytokine production.
Fig 5.
Distinct signatures of TFs in the multiple omics analysis.
(A) Dot plot of the transcriptomic differential distribution of differentially expressed TFs (DETFs) in mouse alveolar macrophages. Horizontal coordinates represent log2(FC) of DEGs, yellow is upregulated, blue is downregulated, and vertical coordinates represent motif-enriched TF significance. (B) Dot plot of the transcriptomic differential distribution of differentially expressed TFs (DETFs) in mouse lung tissues. Horizontal coordinates represent the log2(FC) of DEGs; red indicates upregulation, and green indicates downregulation. (C) Multiset visualization depicting the overlap of TFs shared by lung tissues and cells with DEGs from mouse alveolar macrophages and DEGs from lung tissues. (D) Heatmap depicting the gene expression of DETFs shared by lung tissues and alveolar macrophages. The cell is on the left, and the tissue is on the right. (E) Lollipop plots show transcription factors whose abundance in the ubiquitinated proteome is significantly different in both mouse alveolar cells and tissues. (F) IGV browser view shows multiple omics-altered transcription factors in DARs, with mouse alveolar macrophages in the red area and lung tissues in the blue area.
Fig 6.
Altered downstream signaling molecules of Stat1 and Stat3 upon P. aeruginosa infection.
(A) Network diagram showing the Stat1 and Stat3 target DEGs in the DARs predicted using Homer. (B) Heatmap showing the gene expression of Stat1 and Stat3 target genes found in mouse alveolar macrophages and lung tissue. (C) A schematic model of Stat1- and Stat3-mediated regulation during pulmonary infection with P. aeruginosa. Lung model created with BioRender.com.