The green revolution’s breeding of semi-dwarf rice cultivars in the 1960s improved crop yields, with large increases in the use of nitrogen (N) fertilizer. However, excess N application has caused serious environmental problems, including acid rain and the eutrophication of rivers and oceans. To use N to improve crop yields, while minimizing the associated environmental costs, there is a need to produce crops with higher N-use efficiency and higher yield components. Here we show that transgenic rice overproducing ribulose 1,5-bisphosphate carboxylase–oxygenase (Rubisco)—the key enzyme of photosynthesis—exhibits increased yields with improved N-use efficiency for increasing biomass production when receiving sufficient N fertilization in an experimental paddy field. This field experiment demonstrates an improvement in photosynthesis linked to yield increase due to a higher N-use efficiency in a major crop.

It has been reported that overproduction of Rubisco activase (RCA) in rice (Oryza sativa L.) decreased Rubisco content, resulting in declining photosynthesis. We examined the effects of RCA levels on Rubisco content using transgenic rice with overexpressed or suppressed RCA under the control of different promoters of the RCA and Rubisco small subunit (RBCS) genes. All plants were grown hydroponically with different N concentrations (0.5, 2.0 and 8.0 mM-N). In RCA overproduced plants with > 2-fold RCA content (RCA-HI lines), a 10%–20% decrease in Rubisco content was observed at 0.5 and 2.0 mM-N. In contrast, at 8.0 mM-N, Rubisco content did not change in RCA-HI lines. Conversely, in plants with 50%–60% increased RCA content (RCA-MI lines), Rubisco levels remained unchanged, regardless of N concentration. Such effects on Rubisco content were independent of the promoter that was used. In plants with RCA suppression to < 10% of the wild-type RCA content, Rubisco levels were increased at 0.5 mM-N, but were unchanged at 2.0 and 8.0 mM-N. Thus, the effects of the changes in RCA levels on Rubisco content depended on N supply. Moreover, RCA overproduction was feasible without a decrease in Rubisco content, depending on the degree of RCA production.

Genome-wide association studies (GWASs) are used to detect quantitative trait loci (QTLs) using genomic and phenotypic data as inputs. While genomic data are obtained with high throughput and low cost, obtaining phenotypic data requires a large amount of effort and time. In past breeding programs, researchers and breeders have conducted a large number of phenotypic surveys and accumulated results as legacy data. In this study, we conducted a GWAS using phenotypic data of temperate japonica rice (Oryza sativa) varieties from a public database. The GWAS using the legacy data detected several known agriculturally important genes, indicating reliability of the legacy data for GWAS. By comparing the GWAS using legacy data (L-GWAS) and a GWAS using phenotypic data that we measured (M-GWAS), we detected reliable QTLs for agronomically important traits. These results suggest that a L-GWAS is a strong alternative to replicate tests to confirm the reproducibility of QTLs detected by an M-GWAS. In addition, because legacy data have often been accumulated for many traits, it is possible to evaluate the pleiotropic effect of the QTLs identified for the specific trait that we focused on with respect to various other traits. This study demonstrates the effectiveness of using legacy data for GWASs and proposes the use of legacy data to accelerate genomic breeding.

Overproduction of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco; E.C. 4.1.1.39) in rice did not necessarily improve photosynthesis. The reason for this was that a partial deactivation of Rubisco occurred in RBCS-overexpressed rice plants. Since a negative correlation between the amounts of Rubisco activase (RCA) and Rubisco has been reported for plants with overproduced or decreased RCA, the possibility that RCA content declines in RBCS-overexpressed rice plants was considered. The relationship between RCA and Rubisco contents was examined in RBCS-overexpressed and RBCS-antisense rice plants. Whereas the ratio of RCA to Rubisco contents in RBCS-antisense plants increased three- to fourfold as compared with that of the wild-type levels, this ratio decreased 60–70% of the wild-type levels in RBCS-overexpressed rice plants. Thus, an apparent trade-off between the amounts of RCA and Rubisco was observed in RBCS-transgenic rice plants. However, the amounts of several Calvin–Benson cycle enzymes changed in a similar manner to that of RCA in both types of RBCS-transgenic rice plants. When the relationships between the amounts of these enzymes, including RCA, and those of total leaf-N minus Rubisco-N and trichloroacetic acid (TCA)-soluble N were examined, there were no differences between them irrespective of genotypes. These results indicate that the negative correlation between the amounts of RCA and Rubisco in RBCS-transgenic rice plants is the result of a change in N allocation to Rubisco in transgenic rice plants rather than a trade-off. Such a negative correlation was also found for other Calvin–Benson cycle enzymes. In addition, since the amounts of these Calvin–Benson cycle enzymes and RCA were highly correlated with their mRNA levels irrespective of genotype, it is suggested that changes in the amounts of these proteins are regulated at their transcript levels by a change in N allocation to Rubisco.

The availability of inorganic phosphate (P_i) for ATP synthesis is thought to limit photosynthesis at elevated [CO₂] when P_i regeneration via sucrose or starch synthesis is limited. We report here another mechanism for the occurrence of P_i-limited photosynthesis caused by insufficient capacity of chloroplast triose phosphate isomerase (cpTPI). In cpTPI-antisense transgenic rice (Oryza sativa) plants with 55%–86% reductions in cpTPI content, CO₂ sensitivity of the rate of CO₂ assimilation (A) decreased and even reversed at elevated [CO₂]. The pool sizes of the Calvin–Benson cycle metabolites from pentose phosphates to 3-phosphoglycerate increased at elevated [CO₂], whereas those of ATP decreased. These phenomena are similar to the typical symptoms of P_i-limited photosynthesis, suggesting sufficient capacity of cpTPI is necessary to prevent the occurrence of P_i-limited photosynthesis and that cpTPI content moderately affects photosynthetic capacity at elevated [CO₂]. As there tended to be slight variations in the amounts of total leaf-N depending on the genotypes, relationships between A and the amounts of cpTPI were examined after these parameters were expressed per unit amount of total leaf-N (A/N and cpTPI/N, respectively). A/N at elevated [CO₂] decreased linearly as cpTPI/N decreased before A/N sharply decreased, owing to further decreases in cpTPI/N. Within this linear range, decreases in cpTPI/N by 80% led to decreases up to 27% in A/N at elevated [CO₂]. Thus, cpTPI function is crucial for photosynthesis at elevated [CO₂].

Despite the essentiality of Mn in terrestrial plants, its excessive accumulation in plant tissues can cause growth defects, known as Mn toxicity. Mn toxicity can be classified into apoplastic and symplastic types depending on its onset. Symplastic Mn toxicity is hypothesised to be more critical for growth defects. However, details of the relationship between growth defects and symplastic Mn toxicity remain elusive. In this study, we aimed to elucidate the molecular mechanisms underlying symplastic Mn toxicity in rice plants. We found that under excess Mn conditions, CO₂ assimilation was inhibited by stomatal closure, and both carbon anabolic and catabolic activities were decreased. In addition to stomatal dysfunction, stomatal and leaf anatomical development were also altered by excess Mn accumulation. Furthermore, indole acetic acid (IAA) concentration was decreased, and auxin-responsive gene expression analyses showed IAA-deficient symptoms in leaves due to excess Mn accumulation. These results suggest that excessive Mn accumulation causes IAA deficiency, and low IAA concentrations suppress plant growth by suppressing stomatal opening and leaf anatomical development for efficient CO₂ assimilation in leaves.