OsCBL5-CIPK1-PP23 module enhances rice grain size and weight through the gibberellin pathway.

Grain size is a key factor in determining rice (Oryza sativa) yield, and exploring new pathways to regulate grain size has immense potential to improve yield. In this study, we report that OsCBL5 encodes a calcineurin B subunit protein that significantly promotes grain size and weight. oscbl5 plants produced obviously smaller and lighter seeds. We further revealed that OsCBL5 promotes grain size by affecting cell expansion in the spikelet hull. Biochemical analyses demonstrated that CBL5 interacts with CIPK1 and PP23. Furthermore, double and triple mutations were induced using CRISPR/Cas9 (cr) to analyze the genetic relationship. It was found that the cr-cbl5/cipk1 phenotype was similar to that of cr-cipk1 and that the cr-cbl5/pp23, cr-cipk1/pp23, and cr-cbl5/cipk1/pp23 phenotype was similar to that of cr-pp23, indicating that OsCBL5, CIPK1, and PP23 act as a molecular module influencing seed size. In addition, the results show that both CBL5 and CIPK1 are involved in the gibberellic acid (GA) pathway and significantly affect the accumulation of endogenous active GA4 . PP23 participates in GA signal transduction. In brief, this study identified a new module that affects rice grain size, OsCBL5-CIPK1-PP23, which could potentially be targeted to improve rice yield.

Evaluation of quantitative synchrotron radiation micro-X-ray fluorescence in rice grain.

Concentrations of nutrients and contaminants in rice grain affect human health, specifically through the localization and chemical form of elements. Methods to spatially quantify the concentration and speciation of elements are needed to protect human health and characterize elemental homeostasis in plants. Here, an evaluation was carried out using quantitative synchrotron radiation microprobe X-ray fluorescence (SR-µXRF) imaging by comparing average rice grain concentrations of As, Cu, K, Mn, P, S and Zn measured with rice grain concentrations from acid digestion and ICP-MS analysis for 50 grain samples. Better agreement was found between the two methods for high-Z elements. Regression fits between the two methods allowed quantitative concentration maps of the measured elements. These maps revealed that most elements were concentrated in the bran, although S and Zn permeated into the endosperm. Arsenic was highest in the ovular vascular trace (OVT), with concentrations approaching 100 mg kg-1 in the OVT of a grain from a rice plant grown in As-contaminated soil. Quantitative SR-µXRF is a useful approach for comparison across multiple studies but requires careful consideration of sample preparation and beamline characteristics.

The OsNAC24-OsNAP protein complex activates OsGBSSI and OsSBEI expression to fine-tune starch biosynthesis in rice endosperm.

Starch accounts for up to 90% of the dry weight of rice endosperm and is a key determinant of grain quality. Although starch biosynthesis enzymes have been comprehensively studied, transcriptional regulation of starch-synthesis enzyme-coding genes (SECGs) is largely unknown. In this study, we explored the role of a NAC transcription factor, OsNAC24, in regulating starch biosynthesis in rice. OsNAC24 is highly expressed in developing endosperm. The endosperm of osnac24 mutants is normal in appearance as is starch granule morphology, while total starch content, amylose content, chain length distribution of amylopectin and the physicochemical properties of the starch are changed. In addition, the expression of several SECGs was altered in osnac24 mutant plants. OsNAC24 is a transcriptional activator that targets the promoters of six SECGs; OsGBSSI, OsSBEI, OsAGPS2, OsSSI, OsSSIIIa and OsSSIVb. Since both the mRNA and protein abundances of OsGBSSI and OsSBEI were decreased in the mutants, OsNAC24 functions to regulate starch synthesis mainly through OsGBSSI and OsSBEI. Furthermore, OsNAC24 binds to the newly identified motifs TTGACAA, AGAAGA and ACAAGA as well as the core NAC-binding motif CACG. Another NAC family member, OsNAP, interacts with OsNAC24 and coactivates target gene expression. Loss-of-function of OsNAP led to altered expression in all tested SECGs and reduced the starch content. These results demonstrate that the OsNAC24-OsNAP complex plays key roles in fine-tuning starch synthesis in rice endosperm and further suggest that manipulating the OsNAC24-OsNAP complex regulatory network could be a potential strategy for breeding rice cultivars with improved cooking and eating quality.

Nutrient accumulation and transcriptome patterns during grain development in rice.

Rice is an important source of calories and mineral nutrients for more than half of the world's population. The accumulation of essential and toxic mineral elements in rice grain affects its nutritional quality and safety. However, the patterns and processes by which different elements progressively accumulate during grain filling remain largely unknown. In the present study, we investigated temporal changes in dry matter, elemental concentrations, and the transcriptome in the grain of field-grown rice. We also investigated the effects of seed setting rate and the position of the grain within the rice panicle on element accumulation. Three different patterns of accumulation were observed: (i) elements including K, Mn, B, and Ca showed an early accumulation pattern; (ii) dry matter and elements including N, P, S, Mg, Cu, Zn, Mo, As, and Cd showed a mid accumulation pattern; and (iii) elements such as Fe showed a gradual increase pattern. These different accumulation patterns can be explained by the differences in the biogeochemical behavior of the various elements in the soil, as well as differences in plant nutrient redistribution, gene expression, and the sink-source relationship. These results improve our knowledge of the dynamics of elemental accumulation in rice grain and are helpful for identification of functional genes mediating the translocation of elements to grain.

Low Cd-accumulating rice grain production through inoculation of germinating rice seeds with a plant growth-promoting endophytic bacterium.

This study investigated the effects of the plant growth-promoting endophytic bacterium Cupriavidus taiwanensis KKU2500-3 on the growth of KDML105 rice plants and cadmium (Cd) accumulation in grains. The rice plants were cultivated in soils with 20 and 50 ppm Cd under greenhouse conditions for two consecutive years. At both levels, Cd reduced rice growth and development. Under Cd stress, KKU2500-3 colonized the root surface and interior of rice plants at the early growth stage, and this colonization remained until the late stage. The colonized bacteria increased the pigment contents but reduced the root-to-aboveground translocation of Cd. In soil with 20 ppm Cd, the phytochelatin content of the bacteria-inoculated rice was lower (32.3-89.3%) than that of uninoculated rice. In soil with 50 ppm Cd, the bacteria-inoculated rice exhibited higher glutathione reductase (5-63%) and proline (5-115%) levels, a higher reduced glutathione (GSH)/0.5 oxidized glutathione (GSSG) ratio (4-212%) and decreased lipid peroxidation (1-19%) compared with uninoculated rice. The root-to-grain translocation factor of inoculated rice in soil with 50 ppm Cd was significantly lower than that of inoculated rice in soil with 20 ppm Cd, and this finding was consistent with the 38.6% and 75.1% reductions in Cd accumulation observed in grains from soils with 20 and 50 ppm Cd, respectively. The Cd content of KDML105 grains grown in soil with 50 ppm Cd was 0.36 ppm, which is below the Codex standard for polished rice (0.4 ppm). The levels of available P, Zn, and SO42- also affect Cd availability in soil, and colonized KKU2500-3 showed varying responses to different Cd levels. Thus, bacterial inoculation, the Cd level and soil properties play important roles in Cd accumulation in KDML105 rice grains. The role of C. taiwanensis KKU2500-3 on the production of low-Cd-accumulating rice in paddy fields contaminated with a range of Cd levels should be further investigated.

Rice Grain Detection and Counting Method Based on TCLE-YOLO Model.

Thousand-grain weight is the main parameter for accurately estimating rice yields, and it is an important indicator for variety breeding and cultivation management. The accurate detection and counting of rice grains is an important prerequisite for thousand-grain weight measurements. However, because rice grains are small targets with high overall similarity and different degrees of adhesion, there are still considerable challenges preventing the accurate detection and counting of rice grains during thousand-grain weight measurements. A deep learning model based on a transformer encoder and coordinate attention module was, therefore, designed for detecting and counting rice grains, and named TCLE-YOLO in which YOLOv5 was used as the backbone network. Specifically, to improve the feature representation of the model for small target regions, a coordinate attention (CA) module was introduced into the backbone module of YOLOv5. In addition, another detection head for small targets was designed based on a low-level, high-resolution feature map, and the transformer encoder was applied to the neck module to expand the receptive field of the network and enhance the extraction of key feature of detected targets. This enabled our additional detection head to be more sensitive to rice grains, especially heavily adhesive grains. Finally, EIoU loss was used to further improve accuracy. The experimental results show that, when applied to the self-built rice grain dataset, the precision, recall, and mAP@0.5 of the TCLE-YOLO model were 99.20%, 99.10%, and 99.20%, respectively. Compared with several state-of-the-art models, the proposed TCLE-YOLO model achieves better detection performance. In summary, the rice grain detection method built in this study is suitable for rice grain recognition and counting, and it can provide guidance for accurate thousand-grain weight measurements and the effective evaluation of rice breeding.

The kinase OsSK41/OsGSK5 negatively regulates amylose content in rice endosperm by affecting the interaction between OsEBP89 and OsBP5.

Amylose content (AC) is the main factor determining the palatability, viscosity, transparency, and digestibility of rice (Oryza sativa) grains. AC in rice grains is mainly controlled by different alleles of the Waxy (Wx) gene. The AP2/EREBP transcription factor OsEBP89 interacts with the MYC-like protein OsBP5 to synergistically regulate the expression of Wx. Here, we determined that the GLYCOGEN SYNTHASE KINASE 5 (OsGSK5, also named SHAGGY-like kinase 41 [OsSK41]) inhibits the transcriptional activation activity of OsEBP89 in rice grains during amylose biosynthesis. The loss of OsSK41 function enhanced Wx expression and increased AC in rice grains. By contrast, the loss of function of OsEBP89 reduced Wx expression and decreased AC in rice grains. OsSK41 interacts with OsEBP89 and phosphorylates four of its sites (Thr-28, Thr-30, Ser-238, and Thr-257), which makes OsEBP89 unstable and attenuates its interaction with OsBP5. Wx promoter activity was relatively weak when regulated by the phosphomimic variant OsEBP89E -OsBP5 but relatively strong when regulated by the nonphosphorylatable variant OsEBP89A -OsBP5. Therefore, OsSK41-mediated phosphorylation of OsEBP89 represents an additional layer of complexity in the regulation of amylose biosynthesis during rice grain development. In addition, our findings provide four possible sites for regulating rice grain AC via precise gene editing.

A critical analysis of various post-harvest arsenic removal treatments of rice and their impact on public health due to nutrient loss.

Rice (Oryza sativa L.) is particularly susceptible to arsenic (As) accumulation. Currently, to decrease the level of As accumulated in rice, various post-harvest methods, i.e., polishing, parboiling, pH-dependent soaking, washing, and cooking at different rice-to-water ratios (r/w), are being focused, because it removes significant amount of As from rice grain. Depending upon the rice variety and type, i.e., rough (with husk), husked (without husk/brown), or polished rice, these methods can remove 39-54% As by parboiling, 38-55% by polishing, 37-63% by soaking, and 6-80% by washing and cooking. Infants are highly vulnerable to As exposure; thus, these methods can be helpful for the production of rice-based infant foods. Although concern arises during the use of these methods that apart from decreasing the level of As in rice grain, they also lead to a significant loss of nutrients, such as macro- and micro-elements present in rice. Among these discussed methods, parboiling curtails 5-59%, polishing curtails 6-96%, soaking curtails 33-83%, and washing and cooking in different r/w reduce 8-81% of essential nutrients resulting in 2-90% reduction in contribution to the RDI of these nutrients through rice-based diet. Thus, these post-harvest arsenic removal methods, although reduce arsenic induced health hazard, but may also lead to malnutrition and compromised health in the population based on rice diet. There is a need to explore another way to reduce As from rice without compromising the nutrient availability or to supplement these nutrients through grain enrichment or by introducing additional dietary sources by changing eating habits; however, this may impose an extra economic burden on people.

Effect of Organic Amendments on Cadmium Bioavailability in Soil and its Accumulation in Rice Grain.

A pot trial was conducted during the boro (dry) season to evaluate the impact of six traditional organic amendments (OAs) on the growth of SL-8 rice variety in both agricultural and cadmium (Cd) stressed soil at 2% and 4% application rates. Traditional OAs used in the study were cow dung, mustard oil cake (MOC), rice husk, saw dust, tea leaf and vermi compost (VC). Except for cow dung all other OAs were found to remove 99% of Cd from the aqueous solution, while cow dung removed 95%. Rice grain grown in OA-added soil in all application rates contained less Cd than the control. A 2% application rate was found to be more effective in reducing both Cd bioavailability and Cd in grain. OA application in soil significantly influenced soil pH in all cases. Though both bioavailable Cd in soil and grain Cd were reduced by the OA addition, the Cd uptake tendency of SL-8 rice variety markedly increased because of Cd spiking in soil.

Grain and Leaf Anthocyanin Concentration Varies among Purple Rice Varieties and Growing Condition in Aerated and Flooded Soil.

Anthocyanins are a group of pigments responsible for the red-blue color in plant parts, and have potential for health benefits and pharmaceutical ingredients. To evaluate whether anthocyanin concentrations in five purple rice varieties could be varied by water condition, plants were grown in waterlogged and aerobic (well-drained) soil. Grain anthocyanin concentration and grain yield were measured at maturity, while leaf anthocyanin concentrations were measured at booting and flowering stages. Four varieties grown under the waterlogged condition had 2.0−5.5 times higher grain anthocyanin than in the aerobic condition. There was a positive relationship between grain and leaf anthocyanin at booting in the waterlogged condition (r = 0.90, p < 0.05), while grain and leaf anthocyanin were positively correlated at flowering in both the waterlogged (r = 0.88, p < 0.05) and aerobic (r = 0.97, p < 0.01) conditions. The results suggest that water management should be adopted as a practical agronomic tool for improving the anthocyanin concentration of purple rice for specialist markets, but the specific responses between rice varieties to water management should be carefully considered.