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Figure 1 Monitoring of [Ca2+]cyt in soybean cells challenged with fungal metabolite mixtures. Cells were treated with: the whole culture filtrate of Trichoderma (black trace) or non-inoculated culture medium (grey trace) (a); >3 kDa (black trace) and 2+ transient induced by the >3 kDa metabolites is represented out of scale. In d, the inset shows the [Ca2+]cyt changes induced by the simultaneous application of the metabolite fractions (>3 kDa, black trace;
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Figure 2 [Ca2+]cyt responses of soybean cells to metabolite mixtures secreted by the Trichoderma Δech42 mutant. Cells were treated with >3 kDa (black trace) or Δech42 mutant, grown alone (a) or in the presence of Botrytis (b).
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Figure 3 Detection of intracellular ROS accumulation in soybean cells treated with fungal culture filtrates. Intracellular ROS were detected by H2DCF-DA staining in control cells (Co, a'), in cells treated with >3 kDa (b'-e') and Δ ech42 mutant and Botrytis (Δ ech42+Bo, e'and i'). For each treatment light (a-i) and fluorescence (a'-i') microscope images of the same field are presented. All images were acquired with the same exposition time gauged to the higher fluorescence emission intensity obtained with the Botrytis μm.
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Figure 4 Changes in cell viability in response to fungal culture filtrates. Exponentially growing cells were incubated with >3 kDa (a, black boxes) and Δ ech42) are as in Fig. 3. The 100% value correspond to cells treated for 10 min at 100°C. Data are means ± SD of three independent experiments. Bars labeled with a different letter differ significantly (P
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Figure 5 Effect of the fungal metabolite mixtures on caspase 3-like activity, chromatin condensation and ultrastructure of soybean cells. Panel a and b: caspase 3-like activity in cells treated for 30 min with >3 kDa (a, black boxes) and Δech42) are as in Fig. 3. Data are means ± SD of three independent experiments. Bars labeled with a different letter differ significantly (P μm. Panel d: ultrastructural observations of control cells and cells incubated for 15 min with μm.
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