Figure 1.
Effect of LPS stimualtion to S. cerevisiae BY4742.
A. Methylene blue staining of S. cerevisiae BY4742 cells treated with different concentrations of LPS and observed using light microscopy. The solid bars in the photos indicate 10 µm. The concentrations of LPS were listed on top of the photos. B. Drop test showed that S. cerevisiae BY4742 cells treated with 1.0 mg mg/mL LPS were not dead.
Figure 2.
Number of differentially expressed genes regulated in S. cerevisiae BY4742 cells treated with LPS (LPS+) according to transcriptome analysis.
FDR stands for False Discovery Rate [51]. Log2R represents Log2 of the ratio of the gene expression difference of a gene in “LPS+” samples versus the same gene in “Control” sample.
Figure 3.
Analysis of Gene Ontology and KEGG pathway.
Only the significantly enriched GO categories (A) or KEGG items (B) (p-value ≤0.05) in differentially expressed genes were shown. Blue and red bars represent up-regulated and down-regulated genes in LPS-treated cells compared to control, respectively. The percentage represents the ratio of genes within the genome in each category. The number of genes contained in each item is shown in brackets following the item names.
Figure 4.
PPPI networks generated from the induced genes (A) and the repressed genes (B).
Green nodes indicate proteins encoded by the induced genes, while red nodes represent proteins encoded by the repressed genes. Red lines indicate the interactions among proteins encoded by differentially expressed genes.
Table 1.
Major specific gene ontology categories shared by more than 10 clusters derived from the induced and repressed genes-associated networks.
Figure 5.
Protein phosphorylation network generated from the induced genes and the repressed genes, and its MCODE and CentiScaPe analyses.
Green nodes indicate proteins encoded by the induced genes, while red nodes represent proteins encoded by the repressed genes. Deep colored nodes are associated to genes with |Log2R| ≥1.0 while light colored nodes are associated to genes with 0.5≤ |Log2R| ≤1.0. Yellow nodes indicate Atg1p and Tpk1p in Cluster1 and Cluster 2, respectively. Corresponding centrality values in betweeness and nodes degree were sub graphed according to the importance of selected nodes or bottlenecks. Edges represent interactions between proteins or nodes.
Figure 6.
A. Comparison of transcriptional levels of some key genes in LPS-treated S. cerevisiae BY4742 cells analyzed by RT-PCR and transcriptome, respectively. B. Methylene blue staining of some single gene deletion mutants of S. cerevisiae BY4743 after LPS treatment. According to the transcriptome analysis, genes whose transcriptional level did not significantly change in S. cerevisiae BY4742 after LPS treatment were labeled with star symbol (*); transcriptional levels for the other genes could be found in Table S4.
Figure 7.
Flow cytometry analysis of LPS-treated S. cerevisiae BY4742 cells stained by Annexin-V-FITC and PI.
Untreated cells were used as blank control (Control) and 50% heat-treated dead cells were used as positive control (Heated). The cross gate, in order to show the stained proportions, was set by both blank control and positive control.
Figure 8.
Assessment of mitochondrial membrane potential in LPS-treated S. cerevisiae BY4742 cells, using flow cytometry.
Rh123 was used to stain the untreated (Control), LPS-treated (LPS+) and 100% heat-treated cells (Heated). The result was represented by density banding coloring. Gray dots represent the lowest density, while red dots represent the highest density. The gate set “G” was used to show the differences among the three experiments.
Figure 9.
Detection of ROS accumulation in LPS-treated S. cerevisiae BY4742 cells, using flow cytometry.
H2DCFH-DA was used to stain the untreated (Control) and LPS-treated cells (LPS+). The result was represented by density banding coloring. Gray dots represent the lowest density, while red dots represent the highest density. The gate “E” was set by corresponding dye-unloading samples. FL1-DCF represents 2′,7′-dichlorofluorescein while FL2-H assists for data display. The majority populations (red dots) from both samples shew similar ROS levels.
Figure 10.
Analysis of metacaspase activity of LPS-treated S. cerevisiae BY4742 cells by flow cytometry.
FITC-VAD-FMK was used to detect metacaspace activation. Untreated S. cerevisiae BY4742 cells were used as negative control; 100% heat-treated cells were chosen as positive control. The result was represented by density banding coloring. Gray dots represent the lowest density, while red dots represent the highest density. The gate “D” was set to show the differences among the three types of cells. FL1-FITC represents FITC-VAD-FMK while FS assists for data display.