Fig. 1. Iron deficiency is primary cause of lesion phenotype in osacl-a2 mutants. A and B, Lesion mimic phenotype in individual plant (A) and leaf (B) of wild type (WT) and osacl-a2 at the tillering stage (at 60 d after seed sowing) in the paddy field. Scale bars, 10 cm. C and D, ATP-citrate lyase (ACL) activity (C) and citric acid content (D) in leaves of WT and osacl-a2 at the heading stage. E, Iron (Fe) content in leaves of WT and osacl-a2 at the tillering stage in natural environment. F, Phenotype of WT and osacl-a2 leaves treated with the indicated exogenous Fe3+ concentrations at two weeks after germination. S-Fe3+, Starvation (0 μmol/L Fe3+); N-Fe3+, Normal (50 μmol/L Fe3+); E-Fe3+, Excess (500 μmol/L Fe3+). Scale bar, 2 cm. G, Images of the magnified part of rice stems in WT and osacl-a2 treated with the indicated exogenous Fe3+ concentrations at two weeks after germination. Scale bars, 40 μm. H-J, Citric acid content (H), ACL activity (I), and Fe content (J) in leaves of WT and osacl-a2 treated with the indicated exogenous Fe3+ concentrations at two weeks after germination. K-N, Relative expression levels of genes (K, OsFER2; L, OsIRO2; M, OsYSL2; and N, OsFRDL1) related to Fe3+ absorption and transport in WT and osacl-a2 treated with the indicated exogenous Fe3+ concentrations at two weeks after germination. Ubiquitin (LOC_Os06g46770) was used as the internal control. Data in C-E and H-J are mean ± SE (n = 3). ** indicates significant differences at the 0.01 level by Student’s t-test.In K-N, different lowercase letters above bars indicate significant differences at P < 0.05 by Tukey’s multiple comparisons test.

Fig. 2. OsACL-A2 dependent iron uptake positively affects disease resistance. A, Spray inoculation of wild type (WT) ZH8015 and osacl-a2 (treated with the indicated exogenous iron concentrations at two weeks after germination) with Magnaporthe oryzae strain 14-1. Leaves were photographed at 7 d post-inoculation. Scale bars, 2 cm. S-Fe3+, Starvation (0 μmol/L Fe3+); N-Fe3+, Normal (50 μmol/L Fe3+); E-Fe3+, Excess (500 μmol/L Fe3+). B, Relative fungal growth in the first leaf from the top. C, Number of different lesion types in the leaf. D, Transcript levels of OsACL-A2 in WT leaves with (14-1 strain) and without (Mock) M. oryzae infection. LOC_Os03g50885 was used as the internal control. E and F, ACL activity (E) and citric acid content (F) in WT after M. oryzae infection. Data are mean ± SE (n = 5 in B and C; n = 3 in D-F). Different lowercase letters above bars indicate significant differences at P < 0.05 level by Tukey’s multiple comparisons test.