Fig. 3. Effect of 200 nM HrpW_ea or HrpN_ea on H₂O₂ production and on cytosolic Ca²⁺ level ([Ca²⁺]_cyt) of A. thaliana suspension cells. (A) Time course of H₂O₂ accumulation in the medium of cells treated with 200 nM HrpW_ea, 200 nM HrpN_ea or the corresponding volume of desalting buffer. Half dilution of cell suspension with distilled water triggered hypo-osmotic stress used as positive control. (B) Percentage of increase of H₂O₂ accumulation, at 1 hour, in the medium of cells treated with 200 nM HrpW_ea or HrpN_ea, alone or in combination with 10 µM DPI or by the corresponding volume of desalting buffer. Values are given as a percentage with respect to untreated cells (100%). Values are means of eleven replicates. Error bars represent s.e. (C) Changes in [Ca²⁺]_cyt were measured by using transgenic A. thaliana cells expressing aequorin protein. Cells suspended in culture medium containing 1 mM Ca²⁺, were treated with 200 nM HrpW_ea or HrpN_ea or the corresponding volume of desalting buffer. Hypo-osmotic stress induced by diluting the cell suspension 1:1 with distilled water was used as a positive control. Five independent experiments were carried out, with similar results.