BcMkk1 negatively regulated oxalic acid (OA) biosynthesis via BcRim15.

(A) HPLC profiles of the standard OA and the compound(s) in the culture supernatant of each strain. (B) Quantification of OA production for each strain in (A). The peak area in 38B1 was referred to 1. Values on the bars followed by the same letter are not significantly different at P = 0.05. (C) Comparisons in the expression level of BcOAH among 38B1, ΔBcRim15, ΔBcMkk1-BcRim15 and ΔBcPro40. The expression level in 38B1 was referred to 1. Values on the bars followed by the same letter are not significantly different at P = 0.05. (D) Deletion of BcOAH in the ΔBcMkk1 background resulted in dramatically reduced OA production and activity of EHs. (E) Activity of EHs was measured after grown in CM for 5 days. Values on the bars followed by the same letter are not significantly different at P = 0.05. (F) Mycelial growth inhibition of each strain grown on PDA amended with glucanase in (D). Values on the bars followed by the same letter are not significantly different at P = 0.05. (G) BcBmp3 phosphorylation in each strain. After grown in PDB for 2 days, mycelia of each strain treated with 0.3 mg ml^–1 of CR for 2 h were collected for protein extraction. BcBmp3 and phosphorylated BcBmp3 proteins were detected with the anti-Mpk1 and phospho-p44/42 MAPK antibodies, respectively. The intensities of the western blotting bands were quantified with the programme IMAGE QUANT TL. The intensity of phosphorylated BcBmp3 band for each strain is relative to that of BcBmp3 band. Values on the bars followed by the same letter are not significantly different at P = 0.05. (H) Protoplast amount of each strain after treatment with glucanex at 25°C for 2 h. Bar = 20 μm. Values on the bars followed by the same letter are not significantly different at P = 0.05.