FIG 2 .

Hog1 contributes to the transcriptional response to nitrosative stress. (A) Transcript profiling (RNA sequencing) was performed on three independent replicates cultures of C. albicans HOG1 (RM1000+Clp20) (Table S1) and hog1Δ cells (JC50) exposed to 0 or 2.5 mM DPTA-NONOate for 10 min. In wild-type cells, 321 transcripts displayed statistically significant (>2-fold) increases in level in response to the nitrosative stress. Of these, 205 transcripts (64%) were considered to be Hog1-dependent (red nodes) because they were not induced ≥2-fold by nitrosative stress in hog1Δ cells. The remaining 116 transcripts were considered to be Hog1 independent because they were still induced >2-fold in hog1Δ cells. Using Cytoscape, GO term analysis was performed on these gene subsets, and the outputs were displayed as a gene function network in which the red nodes represent Hog1-dependent genes, the dark grey nodes represent Hog1-independent genes, and the central hubs are colored according to the functional category. For example, blue hubs relate to stress, green hubs relate to metabolism, and the light-gray hub is for genes of unknown function. (B) The levels of intracellular ROS were assayed in C. albicans strains exposed to 0 or 2.5 mM DPTA-NONOate by performing cytometry on DHE-stained cells. Wild type (WT), gray; hog1Δ, red; cta4Δ, green; cap1Δ, yellow. DHE intensity is presented on a log scale, and the mean fluorescent intensity for each cytometry profile is shown. The data shown are representative of three independent experiments. (C) The fold induction of classical nitrosative-stress (YHB1) and oxidative-stress (TRR1) transcripts is shown following exposure of the same C. albicans strains to 2.5 mM DPTA-NONOate for 30 min. Using qRT-PCR, transcript levels were measured relative to the internal ACT1 mRNA control and normalized to the level of that transcript in the absence of the stress. Means and standard deviations are shown for three independent replicate experiments. *, P < 0.05; **, P < 0.01; ***, P < 0.001.