INTRODUCTION
In land plants, phosphorus (P) is one of the essential macro-nutrients
required to maintain their growth and reproduce seeds for the next
generation. P is required in the plant cells as structural constituent
of DNA, RNA, phospholipids, and energy coins (ATP, ADP, and AMP)
(Hawkesford et al., 2012). In addition, P plays an important role in
regulating enzymatic reactions and signaling processes through the
protein phosphorylation/dephosphorylation mechanism (Hawkesford et al.,
2012). Owing to its role in various physiological functions, the
requirement of P in land plants is high followed by the requirements of
nitrogen, potassium, calcium, and magnesium among the 17 essential
mineral nutrients, and generally, the P content corresponds to
approximately 0.2 % of the dry matter of plants (Kirkby, 2012).
Land plants can absorb P only in its inorganic phosphate form (Pi)
present in the soil through their roots or mycorrhizae (Bieleski, 1973).
Land plants possess two kinds of Pi transporters, namely low-affinity
and high-affinity transporters (Furihata et al., 1992; Muchhal et al.,
1996; Kai et al., 1997; Leggewie et al., 1997; Liu et al., 1998), and
their Km values are estimated to be 50-100 µM and 2.5-12.3 µMin planta, respectively, based on the radioactive Pi uptake
experiments (Nussaume et al., 2011). These transporter proteins commonly
harbor 12 trans-membrane-spanning regions, with a large hydrophilic
charged part dividing the protein molecules into two distinct domains
containing of six transmembrane regions in its structure. These
transporter proteins transport one Pi with two to four protons
(H+) into the root against the electrochemical and
concentration gradients across the root surface (Ullrich-Eberius et al.,
1981, 1984; Sakano, 1990; Mimura, 1999; Raghothama, 1999). In this
process, the plasma membrane H+-ATPase contributes to
generating the H+ electrochemical gradient and
maintaining the cytoplasmic pH for Pi/H+ symport
(Ullrich-Eberius et al., 1981, 1984; Mimura, 1999; Raghothama, 1999).
The Pi acquisition strategy in land plants is sophisticated and
well-regulated from the transcriptional- to the post-translational level
(Secco et al., 2012; Gu et al., 2016). Under conditions of low Pi
availability, the expression of Pi transporter genes is activated by
Pi-starvation responsive transcription factors such as PHR
protein-harboring MYB-domain and WRKY-proteins (Rubio et al., 2001; Zhou
et al., 2008; Gu et al., 2016). In addition, microRNA399 and microRNA827
support Pi uptake and accumulation in plants at the post-transcriptional
step (Fujii et al., 2005; Aung et al.,2006; Bari et al., 2006; Chiou et
al., 2006; Franco-Zorrilla et al., 2007; Lin et al., 2010; Secco et al.,
2012; Gu et al., 2016). Simultaneously, a negative feedback system to
suppress the excess Pi uptake is switched on, such as the increase in
class 1 SPX domain-containing proteins, which suppress the expression of
Pi-starvation responsive genes, and the non-protein cording geneIPS1, which acts as a target-mimicry of microRNA399 under Pi
starvation conditions (Franso-Zorrilla et al., 2007; Wang et al., 2009;
Liu et al., 2010; Secco et al., 2012; Puga et al., 2014; Wang et al.,
2014). In contrast, under conditions of adequate Pi availability, the Pi
transporters are actively degraded by an E2 ubiquitin conjugase-related
protein, PHO2, thus leading to the down-regulation of Pi absorption
(Delhaize & Randall, 1995; Dong et al., 1998; Aung et al., 2006; Bari
et al., 2006). After Pi absorption from the rhizosphere, the Pi
homeostasis functions in plant cells (Biddulph et al., 1958; Lee et al.,
1990; Mimura, 1995, 1999). When Pi is sufficiently supplied to the
cells, Pi is stored in vacuoles to maintain its concentration in the
cytosol and organelles such as chloroplasts and mitochondria (Mimura et
al., 1990, 1992, 1996; Pratt et al., 2009). On the other hand, under Pi
deficiency, Pi is exported from the vacuoles to the cytosol and is
preferentially distributed into various organelles for maintaining
cellular physiological functions (Mimura et al., 1990, 1992, 1996; Pratt
et al., 2009).
As mentioned above, although Pi is essential for plant growth, excess Pi
application to plants leads to chlorosis and necrosis in the leaves and
finally withering in whole plants. These phenomena have been recognized
as P toxicity (Rossiter, 1951; Bhatti & Loneragan, 1970; Clarkson &
Scattergood, 1982). To our knowledge, P toxicity has been first reported
in 1917 by John W. Shive (Shive, 1918). He examined the effect of
different levels of P application on soybean growth by the soil and
water culture method, concluding that excess Pi application causes
specific injury in soybean leaves (Shive, 1918). Such P toxicity
symptoms have been observed in the leaves of various land plant such as
rice, wheat, barley and Arabidopsis (Bhatti & Loneragan, 1970;
Aung et al., 2006; Chiou et al., 2006; Wang et al., 2009; Liu et al.,
2010). In general, P toxicity occurs when the Pi content exceeds
approximately 1% of the dry matter of leaves (Bhatti & Loneragan,
1970). P toxicity has been assumed to be caused by zinc (Zn)-deficiency
in the leaves because the Zn content decreases depending on the dosage
of Pi application in plants (Singh et al., 1988; Zhu et al., 2001; Zhang
et al., 2002; Hawkesford et al., 2012). Several hypothetical mechanisms
have been proposed to explain P-induced Zn-deficiency in plants. Some of
the major mechanisms include: 1. the dilution effect of Zn on the tissue
growth stimulated by P application; 2. inhibition of Zn absorption by
root under excess P application through mineral interaction in soil
(Loneragan et al., 1979); 3. suppression of Zn translocation from root
to shoot (Singh et al., 1988); 4. inhibition of Zn acquisition depending
on mycorrhizae under excess P application (Ova et al., 2015). In
contrast, several studies have shown that P toxicity is observed in
plant leaves despite normal Zn accumulation in the leaves (Loneragan et
al., 1979; Cakmak & Marschner, 1987; Ova et al., 2015). Therefore, P
toxicity cannot be explained only by the suppression of Zn acquisition
from soil to leaves. Interestingly, Delhaize and Randall (1995) observed
that light intensity directly modulates the P toxicity symptoms in a
Pi-accumulating Arabidopsis mutant (pho2 ), and limiting
the illumination alleviates P toxicity. This observation implies the
involvement of photosynthesis in the occurrence of P toxicity, but the
detailed molecular mechanisms of P toxicity have not yet been addressed
in land plants.
In this study, we investigated the detailed mechanisms of P toxicity
that cause withering in land plants, especially in the leaves. Firstly,
we examined the effects of Pi application on photosynthesis, and
subsequently, found that high Pi application simultaneously limits
photosynthesis and decreases in the scavenging activity of reactive
oxygen species (ROS), as a result of the lower availability of metals
caused by phytic acid synthesis in leaves. Thereafter we discussed the
detailed whole phenomenon of P toxicity in land plants.