pheatmap function  (RStudio)

Journal: Scientific Reports

Article Title: Dual RNA sequencing of a co-culture model of Pseudomonas aeruginosa and human 2D upper airway organoids

doi: 10.1038/s41598-024-82500-w

Figure Lengend Snippet: Transcriptional response of the epithelium to infection with the different PAO1 strains. ( a ) Schematic representation of the analysis. ( b ) Volcano plot showing the log 2 fold change and -log 10 adjusted p-value of all genes, when comparing the transcriptome of 2D organoids exposed to PAO1 WT or PAO1 ΔQS. ( c ) Volcano plot showing the Log 2 fold change and -log 10 adjusted p-value per gene comparing the transcriptome of 2D organoids exposed to PAO1 WT or unexposed controls. Green indicates differentially expressed genes (DEGs) (log 2 fold change > 1 and adjusted p value < 0.05). ( d ) Gene ontology enrichment analysis showing top 10 categories enriched in 2D organoids exposed to PAO1 WT. Left panel: Healthy organoid line. Right: CF organoid line. ( e ) Gene expression heat map of genes from “Response to lipopolysaccharide” GO term category (GO:0032496). Color code indicates culture condition (co-culture or mono-culture) and PAO1 genotype (WT, Δ pqsA or ΔQS). Heatmaps were plotted using the pheatmap function of DESeq2 79 (version 1.36.0) in Rstudio.

Article Snippet: Heatmaps were plotted using the pheatmap function of DESeq2 79 (version 1.36.0) in Rstudio.

Techniques: Infection, Gene Expression, Co-Culture Assay

Journal: Scientific Reports

Article Title: Dual RNA sequencing of a co-culture model of Pseudomonas aeruginosa and human 2D upper airway organoids

doi: 10.1038/s41598-024-82500-w

Figure Lengend Snippet: Transcriptional response of PAO1 to the presence of airway epithelium. ( a ) Schematic representation of the analysis. ( b ) Volcano plot displaying the Log 2 fold change and –log 10 adjusted p-value of all genes, when comparing the PAO1 transcriptomes of co-culture and bacterial mono-culture samples. Green indicates differentially expressed genes (DEGs) (log 2 fold change > 1 and adjusted p value < 0.05). The number of genes upregulated in co-culture and bacterial mono-culture is indicated. ( c ) Gene ontology enrichment analysis showing top 10 categories enriched in PAO1 exposed to airway epithelium in co-culture. ( d ) Normalized count plots of genes involved in CCR pathway, crc and crcZ . ( e ) KEGG pathway pae00910 plot displaying the log 2 fold change of genes involved in denitrification . DEGs from co-culture vs. bacterial culture mono-culture comparison of PAO1 transcriptomes. ( f ) Normalized count plots of genes involved in P. aeruginosa antibiotic resistance. ( g ) Heat map displaying expression of genes involved in P. aeruginosa T6SS. Genes grouped by H1, H2 or H3 T6SS subtype , . Samples grouped by culture condition. Color code indicates culture condition (Green: bacterial mono-culture; magenta: co-culture) and PAO1 genotype (Dark: WT; middle: Δ pqsA ; light: ΔQS). Heatmaps were plotted using the pheatmap function of DESeq2 79 (version 1.36.0) in Rstudio.

Article Snippet: Heatmaps were plotted using the pheatmap function of DESeq2 79 (version 1.36.0) in Rstudio.

Techniques: Co-Culture Assay, Comparison, Expressing

Journal: Scientific Reports

Article Title: Dual RNA sequencing of a co-culture model of Pseudomonas aeruginosa and human 2D upper airway organoids

doi: 10.1038/s41598-024-82500-w

Figure Lengend Snippet: Epithelial effect on PAO1 QS regulation. ( a ) Gene expression heat map of genes involved in PQS, Las or Rhl QS pathways. ( b ) Volcano plots displaying gene log 2 fold change and –log 10 adjusted p-value when comparing the transcriptomes of WT and Δ pqsA PAO1 to those of ΔQS in co-culture (top) and in pure bacterial cultures (bottom). Venn diagrams display the overlap between genes up (right) and downregulated (left) in the comparisons. ( c ) Gene ontology enrichment analysis showing top 10 categories enriched in genes that are specifically upregulated in co-culture in WT and Δ pqsA PAO1 transcriptomes compared to ΔQS. ( d ) Gene expression heat map showing top 50 co-culture-specific DEGs. Genes are color-coded according to the following categories (Yellow: T6SS; purple: T2SS; green: Leucine metabolism; red: other pathways). ( e ) Gene expression heat map of T6SS eukaryotic and prokaryotic effectors. Sample color code indicates culture condition (Green: bacterial culture mono-culture; magenta: co-culture) and PAO1 genotype (Dark: WT; middle: Δ pqsA ; light: ΔQS). Heatmaps were plotted using the pheatmap function of DESeq2 (version 1.36.0) in Rstudio.

Article Snippet: Heatmaps were plotted using the pheatmap function of DESeq2 79 (version 1.36.0) in Rstudio.

Techniques: Gene Expression, Co-Culture Assay

Journal: Scientific Reports

Article Title: Dual RNA sequencing of a co-culture model of Pseudomonas aeruginosa and human 2D upper airway organoids

doi: 10.1038/s41598-024-82500-w

Figure Lengend Snippet: Benchmarking co-culture model with in vivo P. aeruginosa transcriptomic datasets directly isolated from the airways of CF subjects. ( a ) Accuracy analysis using gene-wise mean and standard deviation values from in vivo samples in Lewin et al., 2023 as reference. Genes were considered accurate if their expression was within 2 standard deviations of the mean in the in vivo sample. ( b ) PCA plots showing sample distribution by condition (Magenta: co-culture; green: bacterial culture in isolates; purple: in vivo) or by study of origin (Orange: this study; purple: Cornforth et al., 2018 ; pink: Kordes et al., 2019 ; blue: Rossi et al., 2018 ). ( c ) Volcano plots displaying gene log 2 fold change and -log 10 adjusted p-value comparing transcriptomes of in vivo P. aeruginosa (left) or co-cultured PAO1 (right) to those of all pure bacterial culture samples. Green indicates differentially expressed genes (DEGs) (log 2 fold change > 1 and adjusted p value < 0.05). Indicated in the boxes the number of up- or downregulated DEGs. ( d ) Venn diagrams displaying the overlap between genes that are upregulated (left), downregulated (middle), or both (right) in the previous in vivo and co-culture comparison to in vitro and mono-culture samples (b). ( e ) Expression heat map displaying the common up-(left) and downregulated (right) genes. Samples clustered based on the expression of all genes plotted per heat map. Color-code indicates condition (Magenta: co-culture; green: pure bacteria; purple: in vivo) and study of origin (Orange: this study; purple: Cornforth et al., 2018; pink: Kordes et al., 2019; blue: Rossi et al., 2018). Heatmaps were plotted using the pheatmap function of DESeq2 (version 1.36.0) in Rstudio. ( f ) Protein-protein interaction network of common DEGs (in vivo and co-culture. Each node represents a protein encoded by a DEG. Edges represent known protein-protein association (either physical or functional) with a confidence level higher than 0.7. Node color represent clusters generated MCL method. Highlighted the pathway to which the cluster proteins belong. ( g ) Log 2 normalized count plots of representative genes from pathways highlighted by the network analysis. Color-code indicates Magenta: co-culture, green: pure bacteria and purple: in vivo.

Article Snippet: Heatmaps were plotted using the pheatmap function of DESeq2 79 (version 1.36.0) in Rstudio.

Techniques: Co-Culture Assay, In Vivo, Isolation, Standard Deviation, Expressing, Cell Culture, Comparison, In Vitro, Bacteria, Functional Assay, Generated

Journal: Scientific Reports

Article Title: Dual RNA sequencing of a co-culture model of Pseudomonas aeruginosa and human 2D upper airway organoids

doi: 10.1038/s41598-024-82500-w

Figure Lengend Snippet: Transcriptional response of the epithelium to infection with the different PAO1 strains. ( a ) Schematic representation of the analysis. ( b ) Volcano plot showing the log 2 fold change and -log 10 adjusted p-value of all genes, when comparing the transcriptome of 2D organoids exposed to PAO1 WT or PAO1 ΔQS. ( c ) Volcano plot showing the Log 2 fold change and -log 10 adjusted p-value per gene comparing the transcriptome of 2D organoids exposed to PAO1 WT or unexposed controls. Green indicates differentially expressed genes (DEGs) (log 2 fold change > 1 and adjusted p value < 0.05). ( d ) Gene ontology enrichment analysis showing top 10 categories enriched in 2D organoids exposed to PAO1 WT. Left panel: Healthy organoid line. Right: CF organoid line. ( e ) Gene expression heat map of genes from “Response to lipopolysaccharide” GO term category (GO:0032496). Color code indicates culture condition (co-culture or mono-culture) and PAO1 genotype (WT, Δ pqsA or ΔQS). Heatmaps were plotted using the pheatmap function of DESeq2 79 (version 1.36.0) in Rstudio.

Article Snippet: Heatmaps were plotted using the pheatmap function of DESeq2 (version 1.36.0) in Rstudio. ( f ) Protein-protein interaction network of common DEGs (in vivo and co-culture.

Techniques: Infection, Gene Expression, Co-Culture Assay

Journal: Scientific Reports

Article Title: Dual RNA sequencing of a co-culture model of Pseudomonas aeruginosa and human 2D upper airway organoids

doi: 10.1038/s41598-024-82500-w

Figure Lengend Snippet: Transcriptional response of PAO1 to the presence of airway epithelium. ( a ) Schematic representation of the analysis. ( b ) Volcano plot displaying the Log 2 fold change and –log 10 adjusted p-value of all genes, when comparing the PAO1 transcriptomes of co-culture and bacterial mono-culture samples. Green indicates differentially expressed genes (DEGs) (log 2 fold change > 1 and adjusted p value < 0.05). The number of genes upregulated in co-culture and bacterial mono-culture is indicated. ( c ) Gene ontology enrichment analysis showing top 10 categories enriched in PAO1 exposed to airway epithelium in co-culture. ( d ) Normalized count plots of genes involved in CCR pathway, crc and crcZ . ( e ) KEGG pathway pae00910 plot displaying the log 2 fold change of genes involved in denitrification . DEGs from co-culture vs. bacterial culture mono-culture comparison of PAO1 transcriptomes. ( f ) Normalized count plots of genes involved in P. aeruginosa antibiotic resistance. ( g ) Heat map displaying expression of genes involved in P. aeruginosa T6SS. Genes grouped by H1, H2 or H3 T6SS subtype , . Samples grouped by culture condition. Color code indicates culture condition (Green: bacterial mono-culture; magenta: co-culture) and PAO1 genotype (Dark: WT; middle: Δ pqsA ; light: ΔQS). Heatmaps were plotted using the pheatmap function of DESeq2 79 (version 1.36.0) in Rstudio.

Article Snippet: Heatmaps were plotted using the pheatmap function of DESeq2 (version 1.36.0) in Rstudio. ( f ) Protein-protein interaction network of common DEGs (in vivo and co-culture.

Techniques: Co-Culture Assay, Comparison, Expressing

Journal: Scientific Reports

Article Title: Dual RNA sequencing of a co-culture model of Pseudomonas aeruginosa and human 2D upper airway organoids

doi: 10.1038/s41598-024-82500-w

Figure Lengend Snippet: Epithelial effect on PAO1 QS regulation. ( a ) Gene expression heat map of genes involved in PQS, Las or Rhl QS pathways. ( b ) Volcano plots displaying gene log 2 fold change and –log 10 adjusted p-value when comparing the transcriptomes of WT and Δ pqsA PAO1 to those of ΔQS in co-culture (top) and in pure bacterial cultures (bottom). Venn diagrams display the overlap between genes up (right) and downregulated (left) in the comparisons. ( c ) Gene ontology enrichment analysis showing top 10 categories enriched in genes that are specifically upregulated in co-culture in WT and Δ pqsA PAO1 transcriptomes compared to ΔQS. ( d ) Gene expression heat map showing top 50 co-culture-specific DEGs. Genes are color-coded according to the following categories (Yellow: T6SS; purple: T2SS; green: Leucine metabolism; red: other pathways). ( e ) Gene expression heat map of T6SS eukaryotic and prokaryotic effectors. Sample color code indicates culture condition (Green: bacterial culture mono-culture; magenta: co-culture) and PAO1 genotype (Dark: WT; middle: Δ pqsA ; light: ΔQS). Heatmaps were plotted using the pheatmap function of DESeq2 (version 1.36.0) in Rstudio.

Article Snippet: Heatmaps were plotted using the pheatmap function of DESeq2 (version 1.36.0) in Rstudio. ( f ) Protein-protein interaction network of common DEGs (in vivo and co-culture.

Techniques: Gene Expression, Co-Culture Assay

Journal: Scientific Reports

Article Title: Dual RNA sequencing of a co-culture model of Pseudomonas aeruginosa and human 2D upper airway organoids

doi: 10.1038/s41598-024-82500-w

Figure Lengend Snippet: Benchmarking co-culture model with in vivo P. aeruginosa transcriptomic datasets directly isolated from the airways of CF subjects. ( a ) Accuracy analysis using gene-wise mean and standard deviation values from in vivo samples in Lewin et al., 2023 as reference. Genes were considered accurate if their expression was within 2 standard deviations of the mean in the in vivo sample. ( b ) PCA plots showing sample distribution by condition (Magenta: co-culture; green: bacterial culture in isolates; purple: in vivo) or by study of origin (Orange: this study; purple: Cornforth et al., 2018 ; pink: Kordes et al., 2019 ; blue: Rossi et al., 2018 ). ( c ) Volcano plots displaying gene log 2 fold change and -log 10 adjusted p-value comparing transcriptomes of in vivo P. aeruginosa (left) or co-cultured PAO1 (right) to those of all pure bacterial culture samples. Green indicates differentially expressed genes (DEGs) (log 2 fold change > 1 and adjusted p value < 0.05). Indicated in the boxes the number of up- or downregulated DEGs. ( d ) Venn diagrams displaying the overlap between genes that are upregulated (left), downregulated (middle), or both (right) in the previous in vivo and co-culture comparison to in vitro and mono-culture samples (b). ( e ) Expression heat map displaying the common up-(left) and downregulated (right) genes. Samples clustered based on the expression of all genes plotted per heat map. Color-code indicates condition (Magenta: co-culture; green: pure bacteria; purple: in vivo) and study of origin (Orange: this study; purple: Cornforth et al., 2018; pink: Kordes et al., 2019; blue: Rossi et al., 2018). Heatmaps were plotted using the pheatmap function of DESeq2 (version 1.36.0) in Rstudio. ( f ) Protein-protein interaction network of common DEGs (in vivo and co-culture. Each node represents a protein encoded by a DEG. Edges represent known protein-protein association (either physical or functional) with a confidence level higher than 0.7. Node color represent clusters generated MCL method. Highlighted the pathway to which the cluster proteins belong. ( g ) Log 2 normalized count plots of representative genes from pathways highlighted by the network analysis. Color-code indicates Magenta: co-culture, green: pure bacteria and purple: in vivo.

Article Snippet: Heatmaps were plotted using the pheatmap function of DESeq2 (version 1.36.0) in Rstudio. ( f ) Protein-protein interaction network of common DEGs (in vivo and co-culture.

Techniques: Co-Culture Assay, In Vivo, Isolation, Standard Deviation, Expressing, Cell Culture, Comparison, In Vitro, Bacteria, Functional Assay, Generated