The total leaf N content significantly decreased in the low-Pi
treatments and remained almost
constant
above 1.2 mM Pi treatments (Figure 4a). The Rubisco content showed a
similar response to the total leaf N content. A low-Pi application
slightly decreased the Rubisco content, but a higher Pi application did
not change the Rubisco content (Figure 4b).
Rubisco activation maintained high in the low-Pi treatments, but
decreased with increasing Pi application (Figure 4c). The carbamylation
potential of Rubisco, which is lowered by a tight binging inhibitor,
such as 2-carboxy-D-arabinitol 1-phosphate, was not different among
different levels of Pi application (Figure 4d). Indeed, the Rubisco in
the night-sampled leaves showed significantly lower activation and
carbamylation potential compared with the Rubisco sampled under
illumination (Figure 4c, d). Thus, one of the reasons for the decline in
photosynthesis by excessive Pi application is a decline in in
vivo Rubisco activation.
Because Rubisco activation is catalyzed by Rubisco activase (RCA)
(Salvucci et al., 1985), we examined the RCA content of the leaves.
Three different isoforms of RCA resulting from alternative splicing and
limited proteolysis have been reported in rice plants (Portis 2003;
Vargas-Suarez et al., 2004; Fukayama et al., 2012). According to
Fukayama et al. (2012), an upper protein band corresponds to theα- form of RCA, which has redox-active cysteine residues; a middle
protein band corresponds to the β -form, which lacks redox active
Cys residues; and a lower protein band corresponds to the N-terminal
processed β (β* )-form in the leaves (Figure 4e). The total
amount of RCA, including the α- , β-, and β*- forms
was similar among different Pi treatments (Figure 4f). However, a
high-Pi application decreased the content of α - andβ -forms compared with the control-Pi conditions (Figure 4g).
Contrary to the change in the α- and β -forms, theβ* -form content increased with increasing Pi application (Figure
4g). When the isoform ratios of α /β and β */βwere calculated, an increase in Pi application decreased theα /β ratio and increased the β */β ratio,
compared to the low-Pi and control plants (Figure 4h, i). These results
indicated that an increase in Pi application modified the RCA isoform
composition of the leaves.