Significance Phosphate (Pi) is a primary nutrient for plant growth. Because of the low availability of soil Pi, the Pi starvation signaling in plants is gaining great interest. Arabidopsis AtPHR1 and its rice homologue OsPHR2 are known to be central transcription factors in Pi homeostasis; however, the mechanism of how plants sense external Pi fluctuation to regulate the activity of AtPHR1/OsPHR2 has been elusive. Here, we identify rice SPX1 and SPX2 as Pi-dependent inhibitors of PHR2, implicating SPX1 and SPX2 in the Pi-sensing mechanism. We also show that the SPX domain of SPX1 and SPX2 is critical for repressing PHR2 binding to cis elements by protein interaction. The discovery of cellular nutrient concentration-dependent fine-tuning sheds light on a novel mechanism of plant adaption to environmental cues.
Abstract Photorespiration, often considered as a wasteful process, is a key target for bioengineering to improve crop yields. Several photorespiratory bypasses have been designed to efficiently metabolize 2-phosphoglycolate and increase the CO2 concentration in chloroplasts, thereby reducing photorespiration. However, the suppression of primary nitrate assimilation remains an issue when photorespiration is inhibited. In this study, we designed a carbon and nitrogen metabolism-coupled photorespiratory bypass, termed the GCBG bypass, in rice (Oryza sativa) chloroplasts. Our results demonstrated efficient assembly and expression of the GCBG bypass in rice chloroplasts, which affected the levels of typical metabolites and their derivatives of natural photorespiration and enhanced the photosynthetic efficiency. Metabolomic analyses revealed that oxaloacetate, produced from glycolate in chloroplasts, positively impacted amino acid synthesis, energy metabolism, and sugar synthesis. The engineered GCBG plants showed an average yield increase of 19.0% (17.8-20.2%) compared to wild-type plants under natural growth conditions, alongside improved nitrogen uptake, which compensated for 44.1% of yield losses under nitrogen-limited conditions. In summary, the GCBG bypass substantially improved the photosynthetic efficiency, biomass and yield in rice by integrating carbon and nitrogen metabolism. This study introduces a strategy for engineering high-yielding rice or other crops with improved photosynthetic efficiency and nitrogen uptake.
Summary Nitrogen (N) is one of the key essential macronutrients that affects rice growth and yield. Inorganic N fertilizers are excessively used to boost yield and generate serious collateral environmental pollution. Therefore, improving crop N use efficiency (NUE) is highly desirable and has been a major endeavour in crop improvement. However, only a few regulators have been identified that can be used to improve NUE in rice to date. Here we show that the rice NIN‐like protein 4 (OsNLP4) significantly improves the rice NUE and yield. Field trials consistently showed that loss‐of‐ OsNLP4 dramatically reduced yield and NUE compared with wild type under different N regimes. In contrast, the OsNLP4 overexpression lines remarkably increased yield by 30% and NUE by 47% under moderate N level compared with wild type. Transcriptomic analyses revealed that OsNLP4 orchestrates the expression of a majority of known N uptake, assimilation and signalling genes by directly binding to the nitrate‐responsive cis ‐element in their promoters to regulate their expression. Moreover, overexpression of OsNLP4 can recover the phenotype of Arabidopsis nlp7 mutant and enhance its biomass. Our results demonstrate that OsNLP4 plays a pivotal role in rice NUE and sheds light on crop NUE improvement.
Abstract Auxin is a phytohormone that plays an important role in plant growth and development by forming local concentration gradients. The regulation of auxin levels is determined by the activity of auxin efflux carrier protein PIN-formed (PIN). In Arabidopsis thaliana, PIN-formed1 (PIN1) functions in inflorescence and root development. In rice (Oryza sativa L.), there are four PIN1 homologs (OsPIN1a–1d), but their functions remain largely unexplored. Hence, in this study, we created mutant alleles of PIN1 gene—pin1a, pin1b, pin1c, pin1d, pin1a pin1b and pin1c pin1d— using CRISPR/Cas9 technology and used them to study the functions of the four OsPIN1 paralogs in rice. In wild-type rice, all four OsPIN1 genes were relatively highly expressed in the root than in other tissues. Compared with the wild type, the OsPIN1 single mutants had no dramatic phenotypes, but the pin1a pin1b double mutant had shorter shoots and primary roots, fewer crown roots, reduced root gravitropism, longer root hairs and larger panicle branch angle. Furthermore, the pin1c pin1d double mutant showed no observable phenotype at the seedling stage, but showed naked, pin-shape inflorescence at flowering. These data suggest that OsPIN1a and OsPIN1b are involved in root, shoot and inflorescence development in rice, whereas OsPIN1c and OsPIN1d mainly function in panicle formation. Our study provides basic knowledge that will facilitate the study of auxin transport and signaling in rice.
Summary The degree of rice tillering is an important agronomic trait that can be markedly affected by nitrogen supply. However, less is known about how nitrogen‐regulated rice tillering is related to polar auxin transport. Compared with nitrate, ammonium induced tiller development and was paralleled with increased 3 H‐indole‐acetic acid (IAA) transport and greater auxin into the junctions. OsPIN9 , an auxin efflux carrier, was selected as the candidate gene involved in ammonium‐regulated tillering based on GeneChip data. Compared with wild‐type plants, ospin9 mutants had fewer tillers, and OsPIN9 overexpression increased the tiller number. Additionally, OsPIN9 was mainly expressed in vascular tissue of the junction and tiller buds, and encoded a membrane‐localised protein. Heterologous expression in Xenopus oocytes and yeast demonstrated that OsPIN9 is a functional auxin efflux transporter. More importantly, its RNA and protein levels were induced by ammonium but not by nitrate, and tiller numbers in mutants did not respond to nitrogen forms. Further advantages, including increased tiller number and grain yield, were observed in overexpression lines grown in the paddy field at a low‐nitrogen rate compared with at a high‐nitrogen rate. Our data revealed that ammonium supply and an auxin efflux transporter co‐ordinately control tiller bud elongation in rice.
Leaf-form ferredoxin-NADP+ oxidoreductases (LFNRs) function in the last step of the photosynthetic electron transport chain, exist as soluble proteins in the chloroplast stroma and are weakly associated with thylakoids or tightly anchored to chloroplast membranes. Arabidopsis thaliana has two LFNRs, and the chloroplast proteins AtTROL and AtTIC62 participate in anchoring AtLFNRs to the thylakoid membrane. By contrast, the membrane anchoring mechanism of rice (Oryza sativa) LFNRs has not been elucidated. Here, we investigated the membrane-anchoring mechanism of LFNRs and its physiological roles in rice. We characterized the rice protein OsTROL1 based on its homology to AtTROL. We determined that OsTROL1 is also a thylakoid membrane anchor and its loss leads to a compensatory increase in OsTIC62. OsLFNR1 attachment through a membrane anchor depends on OsLFNR2, unlike the Arabidopsis counterparts. In addition, OsTIC62 was more highly expressed in the dark than under light conditions, consistent with the increased membrane binding of OsLFNR in the dark. Moreover, we observed reciprocal stabilization between OsLFNRs and their membrane anchors. In addition, unlike in Arabidopsis, the loss of LFNR membrane anchor affects photosynthesis in rice. Overall, our study sheds light on the mechanisms anchoring LFNRs to membranes in rice and highlights differences with Arabidopsis.
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