To investigate why Cu/Zn-SOD activity decreased in the higher-Pi treatments, the content of several essential nutrient elements was determined using the inductively coupled plasma atomic emission spectroscopy (ICP-AES). The P content increased with increasing Pi application (Figure 7a). The content values of K, Ca, and Fe were not affected by Pi application (Figure 7a), but those of Mg, Mn, Zn, and Cu increased with increasing Pi application (Figure 7a).
To deduce nutrient availabilities within leaves, we next examined the mRNA expressions of several metal deficiency-responsive genes, such asZIP4 (Ishimaru et al., 2005), IRO2 (Ogo et al., 2006) andCOPT1 (Yuan et al., 2011), which are expressed under Zn-, Fe-, and Cu-deficient conditions, respectively. The ZIP4 andIRO2 expressions increased with increasing in Pi application (Figure 7b), COPT1 expression showed no significant change with varying Pi application conditions (Figure 7b). These results suggest that excessive Pi application does not inhibit the metal transport to leaves, but Zn- and Fe-deficiency occurs within cells under conditions of excessive Pi application.