Fig. 3

prxA suppression is responsible for the impairment of fungal resistance to H₂O₂ in G6PD-overexpression strains. All cultivations used NO₃⁻ as the sole nitrogen source. A, B Quantification analysis of intracellular O₂^·− and H₂O₂ in WT (WT_pyrG) and nP.gsdA. After precultivation, both strains were exposed to 0 or 1 mM H₂O₂ for 30 min followed by addition of ROS fluorescent probes. The ROS scavenger NAC (10 mM) was applied 1 h before the probe incubation. Fluorescence intensities of BES-So-AM and BES-H₂O₂-Ac were used to measure the level of intracellular superoxide and H₂O₂, respectively. All values were normalized by that in the unstressed WT (set to 100) (mean ± SD; n = 3, *P < 0.05, **P < 0.001, one-way ANOVA). C Relative expression levels of prxA in WT (WT_pyrG) and nP.gsdA. Strains were precultivated for 16 h, and then exposed to 1 mM H₂O₂ for 30 min. The level of prxA in WT without H₂O₂-treatment was set to 1 (mean ± SD; n = 3, *P < 0.05, one-way ANOVA). D Relative Prx-GFP levels in the WT (P_Gfp) and nP.gsdA (nP.gsdA/P_Gfp) strains. Inset, fluorescence spectra of Prx-GFPs from the corresponding cell lysates. After preculture, both strains were exposed to 0 and 1 mM H₂O₂ for 2 h. Cell lysates (1 mg/ml) were used for fluorescence analysis. E Effects of constitutive expression of prxA on fungal oxidative resistance. Conidia (1 × 10⁵) of the strains were spotted and cultivated for 2 days on NO₃⁻-MM plates with or without 2 mM H₂O₂. Newly constructed strains are as follows: gP.prxA (replacing prxA promoter with gpdA promoter) and nP.gsdA/gP.prxA (replacing gsdA promoter with niaD promoter in gP.prxA)