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CO2-Responsive CCT Protein (CRCT) is suggested to be a positive regulator of starch synthesis-related genes in the vegetative organs of rice. To investigate the physiological function of CRCT, we produced CRCT knockout lines by CRISPR/Cas9 system. CRCT knockout tended to slightly reduce photosynthetic performance and had some adverse effects on growth parameters such as tiller number and straw dry weight. The starch content in the leaf sheath of CRCT knockout lines was significantly lower than that of wild-type. qRT-PCR and RNA-seq analysis showed that sucrose treatment induced the expression of CRCT, which, in turn, induced many starch synthesis-related genes in wild-type, whereas this induction did not occur in CRCT knockout lines. In conclusion, our results suggest that sugar induction of starch synthesis-related genes is almost fully dependent on CRCT, making it a necessary factor for normal photosynthesis and growth of rice. [ABSTRACT FROM AUTHOR]

Rice is the model C3 crop for investigating the starch biosynthesis mechanism in endosperm because of its importance in grain production. However, little is known about starch biosynthesis in the vegetative organs of rice. In this study, we used novel rice mutants by inserting Tos17 into the starch synthase (SS) IIIb gene, which is mainly expressed in the leaf sheath (LS) and leaf blade (LB), and an ss1 mutant to clarify the differences in roles among SS isozymes during starch biosynthesis. Native polyacrylamide gel electrophoresis (PAGE)/activity staining for SS, using LS and LB of ss mutants, revealed that the lowest migrating SS activity bands on the gel were derived from SSIIIb activity and those of two ss3b mutants were not detected. The apparent amylose content of LS starch of ss3b mutants increased. Moreover, the chain-length distribution and size-exclusion chromatography analysis using ss mutants showed that SSIIIb and SSI synthesize the B2–B3 chain and A–B1 chain of amylopectin in the LS and LB respectively. Interestingly, we also found that starch contents were decreased in the LS and LB of ss3b mutants, although SSI deficiency did not affect the starch levels. All these results indicated that SSIIIb synthesizes the long chain of amylopectin in the LS and LB similar to SSIIIa in the endosperm, while SSI synthesizes the short chain in the vegetative organ as the same in the endosperm. [ABSTRACT FROM AUTHOR]

We screened 80 Asian rice (Oryza sativa L.) cultivars for the presence of the submergence-tolerance gene SUB1A-1 and the floating genes SNORKEL1 (SK1) and SNORKEL2 (SK2), and found that the deepwater rice cultivar Baisbish (BSB) and the submergence-tolerant cultivar Flood Resistant 13A (FR13A) both possess the SUB1A-1 and the SK1/2. When BSB and FR13A seedlings were completely submerged, spindly growth of shoots was induced in BSB but not in FR13A. Submergence significantly increased the SUB1A-1 transcript abundance in BSB and FR13A shoots, but the expression level in BSB was much lower than that of FR13A. Submergence also induced the expression of both ERF66 and ERF67, the transcriptional targets of SUB1A-1, in FR13A shoots, whereas it upregulated the expression of ERF67 but not that of ERF66 in BSB shoots. These results suggest that BSB could not display submergence tolerance due to the low expression of SUB1A-1 and/or ERF66 under submergence. [ABSTRACT FROM AUTHOR]

CO2‐responsive CCT protein (CRCT) is a positive regulator of starch synthesis–related genes such as ADP‐glucose pyrophosphorylase large subunit 1 and starch branching enzyme I particularly in the leaf sheath of rice (Oryza sativa L.). The promoter GUS analysis revealed that CRCT expressed exclusively in the vascular bundle, whereas starch synthesis–related genes were expressed in different sites such as mesophyll cell and starch storage parenchyma cell. However, the chromatin immunoprecipitation (ChIP) using a FLAG‐CRCT overexpression line and subsequent qPCR analyses showed that the 5′‐flanking regions of these starch synthesis–related genes tended to be enriched by ChIP, suggesting that CRCT can bind to the promoter regions of these genes. The monomer of CRCT is 34.2 kDa; however, CRCT was detected at 270 kDa via gel filtration chromatography, suggesting that CRCT forms a complex in vivo. Immunoprecipitation and subsequent MS analysis pulled down several 14‐3‐3‐like proteins. A yeast two‐hybrid analysis and bimolecular fluorescence complementation assays confirmed the interaction between CRCT and 14‐3‐3‐like proteins. Although there is an inconsistency in the place of expression, this study provides important findings regarding the molecular function of CRCT to control the expression of key starch synthesis–related genes. CO2‐responsive CCT protein (CRCT) is a positive regulator of starch synthesis in vegetative organs of rice. Here we show that CRCT can interact with 14‐3‐3 proteins and controls the transcription of key starch synthesis–related genes such as ADP‐glucose pyrophosphorylase large subunit 1. [ABSTRACT FROM AUTHOR]

Rubisco is the key enzyme responsible for photosynthetic CO2 fixation. Previously, we succeeded to increase the catalytic rate (kcat) of Rubisco by the overexpression of sorghum C4-type Rubisco small subunit (RbcS) in rice. These Rubisco are present as a chimera of sorghum RbcS and rice RbcS. In this study, we introduced an RNAi construct for rice RbcS into a sorghum RbcS overexpression line (SS10). Obtained double-transgenic lines iSS4 and iSS5 dominantly expressed sorghum RbcS and accumulated 35% and 20% of Rubisco content of non-transgenic rice (WT), respectively. Rubisco in these double-transgenic lines showed higher kcat and Km for CO2 than WT and SS10. Moreover, iSS4 showed higher photosynthetic nitrogen use efficiency under high CO2 conditions. These results demonstrate the usefulness of C4-type RbcS to improve photosynthetic performance under elevated CO2 conditions. [ABSTRACT FROM AUTHOR]