Under
P toxicity conditions, leaf withering would be caused by the oxidative
stress resulting from the disruption of Cu/Zn-SOD activities as well as
photosynthetic limitation. When PET activity exceeds the electron sink
capacity, ROS production is stimulated in both PSII and PSI
(Krieger-Liszkay, 2005; Sonoike, 2011; Pospíšil, 2016; Takagi et al.,
2016a). ROS show high reactivity to biomolecules including DNA, proteins
and lipids; therefore, the accumulation of ROS triggers cellular
dysfunction to cause cell death (Apel & Hirt, 2004). Under high-Pi
conditions, an increase was observed in the values of Y(NO) and Y(NA),
which indicate the reduced states of PSII and PSI, respectively, despite
an increase in the values of NPQ and Y(ND), which contribute to the
suppression of ROS production both in PSII and PSI (Figure 2; Figures
S1) (Müller et al., 2001; Takagi et al., 2017). Wada et al. (2018)
reported that the Rubisco-antisense plants showed an over-reduction of
the PET chain. Furthermore, Takagi et al. (2016b) reported that the
suppression of electron sink activities inhibits both PET reaction andpmf generation. At the same time, the values of
gH+ andV H+ also decreased because of
the decrease in the ATP requirement of electron sink activities (Takagi
et al., 2016b). Here, we observed that the ATP/ADP ratio significantly
increased with increaseing Pi application (Figure 3d). Moreover,pmf , gH+, andV H+ decreased under 3.0 mM Pi
conditions (Figure S2). Therefore, the high Pi accumulation would cause
an over-reduction of the PET chain owing to a decrease in electron sink
activities, including Rubisco deactivation. An increase in APX activity
reconciles with the stimulation of oxidative stress under P toxicity
conditions because the chloroplastic APX activity responds to the
oxidative stress (Figure 5a, b) (Cakmak, 2005). Furthermore, a decrease
in the Chl content which with the decreasing in PSI (Figure 1c; Figure
S4), and the decrease in Fv/Fm supports the oxidative stress on the
thylakoid membranes under excessive Pi application conditions (Figure
2j) (Terashima et al., 1994; Sonoike, 2011; Takagi et al., 2016a). In
addition, we found an increase in the expression of an ROS-responsive
gene PR5 (Ganesan
&
Thomas, 2001), and lipid hydroperoxide content (LOOH) under high-Pi
application conditions (Figure S5a, c). In contrast, the expression of a
programmed cell death-related gene VPE2 (Deng et al., 2011)
decreased under high-Pi conditions (Figure S5b). Based on the
illumination dependency of P toxicity symptoms (Delhaize & Randall,
1995), necrosis but not programmed cell death would be triggered by ROS
production in thylakoid membranes, and this would be intensified by both
the suppression of Cu/Zn-SOD activity and a decrease in
photosynthetic
electron sink capacities (Figure
9).
In the present study, we discussed a detailed mechanism for P toxicity
in rice plants (Figure 9). To date, much attention has been paid to
phytic acid synthesis in seeds. However, our results indicated that to
maintain a proper leaf phytic acid content is important for maintaining
plant growth and escaping oxidative stress triggered by PET reaction.
Because the phytic acid synthesis pathway would be activated by an
increase in the cytosolic Pi content through an increase in the
sugar-phosphate content, the fine-tuning of Pi compartmentation within
cells or modulation of sugar-phosphate metabolic flux might contribute
to improving the Pi-use efficiency in the absence of phytic acid
synthesis and P toxicity symptoms. The present study would open new
opportunity to design breeding strategies for improving P use efficiency
in crop plants. To modify the phytic acid content in land plants is also
an important goal to improve human health because of severe Zn and Fe
deficiency of humans in the world (Perera et al., 2018). That is, our
findings would contribute to minimizing the phytic acid content as well
as maximizing the plant yield for improving human health, especially for
overcoming Zn-deficiency in humans and for fulfilling food demands in
the future. Based on the present knowledge, we must continue to
challenge ourselves to achieve these two important subjects in the main
crop.