水稻遗传育种合辑Rice Genetics · Breeding · Germplasm Resources
OsMas1, a novel maspardin protein gene, confers tolerance to salt and drought stresses by regulating ABA signaling in rice
Drought and salt stresses, the major environmental abiotic stresses in agriculture worldwide, affect plant growth, crop productivity, and quality. Therefore, developing crops with higher drought and salt tolerance is highly desirable. This study reported the isolation, biological function, and molecular characterization of a novel maspardin gene, OsMas1, from rice. The OsMas1 protein was localized to the cytoplasm. The expression levels of OsMas1 were up-regulated under mannitol, PEG6000, NaCl, and abscisic acid (ABA) treatments in rice. The OsMas1 gene was introduced into the rice cultivar Zhonghua 11 (wild type, WT). OsMas1-overexpression (OsMas1-OE) plants exhibited significantly enhanced salt and drought tolerance; in contrast, OsMas1-interference (OsMas1-RNAi) plants exhibited decreased tolerance to salt and drought stresses, compared with WT. OsMas1-OE plants exhibited enhanced hypersensitivity, while OsMas1-RNAi plants showed less sensitivity to exogenous ABA treatment at both germination and post-germination stages. ABA, proline and K+ contents and superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), and photosynthesis activities were significantly increased. In contrast, malonaldehyde (MDA), hydrogen peroxide (H2O2), superoxide anion radical (O2-·), and Na+ contents were significantly decreased in OsMas1-OE plants compared with OsMas1-RNAi and WT plants. Overexpression of OsMas1 up-regulated the genes involved in ABA signaling, proline biosynthesis, reactive oxygen species (ROS)-scavenging system, photosynthesis, and ion transport under salt and drought stresses. Our results indicate that the OsMas1 gene improves salt and drought tolerance in rice, which may serve as a candidate gene for enhancing crop resistance to abiotic stresses.
Rice direct seeding has the significant potential to save labor and water, conserve environmental resources, and reduce greenhouse gas emissions tremendously. Therefore, rice direct seeding is becoming the major cultivation technology applied to rice production in many countries. Identifying and utilizing genes controlling mesocotyl elongation is an effective approach to accelerate breeding procedures and meet the requirements for direct-seeded rice (DSR) production. This study used a permanent mapping population with 144 recombinant inbred lines (RILs) and 2 828 bin-markers to detect quantitative trait loci (QTLs) associated with mesocotyl length in 2019 and 2020. The mesocotyl lengths of the rice RILs and their parents, Lijiangxintuanheigu (LTH) and Shennong 265 (SN265), were measured in a growth chamber at 30°C in a dark environment. A total of 16 QTLs for mesocotyl length were identified on chromosomes 1(2), 2(4), 3(2), 4, 5, 6, 7, 9, 11(2), and 12. Seven of these QTLs, including qML1a, qML1b, qML2d, qML3a, qML3b, qML5, and qML11b, were reproducibly detected in both years via the interval mapping method. The major QTL, qML3a, was reidentified in two years via the composite interval mapping method. A total of 10 to 413 annotated genes for each QTL were identified in their smallest genetic intervals of 37.69 kb to 2.78 Mb, respectively. Thirteen predicted genes within a relatively small genetic interval (88.18 kb) of the major mesocotyl elongation QTL, qML3a, were more thoroughly analyzed. Finally, the coding DNA sequence variations among SN265, LTH, and Nipponbare indicated that the LOC_Os03g50550 gene was the strongest candidate gene for the qML3a QTL controlling the mesocotyl elongation. This LOC_Os03g50550 gene encodes a mitogen-activated protein kinase. Relative gene expression analysis using qRT-RCR further revealed that the expression levels of the LOC_Os03g50550 gene in the mesocotyl of LTH were significantly lower than in the mesocotyl of SN265. In conclusion, these results further strengthen our knowledge about rice’s genetic mechanisms of mesocotyl elongation. This investigation’s discoveries will help to accelerate breeding programs for new DSR variety development.
Rice is the staple food for about half of the world’s population. Preferred by consumers, aromatic rice is a special type of rice with great commercial value. Cooking and eating qualities and aroma are the major grain qualities favored by most consumers. Currently, most of the available aromatic varieties have low yields and some undesirable agronomic traits. Thus, there is an urgent need to develop better aromatic rice varieties. This work aims to identify rice germplasm lines that have good grain quality and to develop new varieties with desirable traits. Thirty-six out of 188 germplasm lines were found to have betaine aldehyde dehydrogenase 2 (badh2) controlling the aroma and were analyzed for their 2-acetyl-1-pyrroline (2AP) contents. Then, 17 of those lines were found to have alleles for low amylose content and low gelatinization temperature, controlled by waxy and starch synthase IIa (SSIIa), respectively, suggesting that they are aromatic rice lines with high cooking and eating qualities. A total of 158 F7 recombinant inbred lines (RILs) generated from five crosses of the selected germplasm lines were planted for phenotypic and yield observations, resulting in 27 F8 RILs selected for yield evaluation and genotyping. Finally, four out of the seven F9 aromatic RILs showed high yield, high 2AP production, and low amylose content, in agreement with their genotypes. The other three F9 RILs were aromatic rice lines with high amylose content and high yield. Because consumer preferences for grain quality vary depending on regions and ethnic groups, the high-yielding aromatic RILs generated from this study can be used to increase the yield of Thai rice and to raise market value and farm profits.
Plant chlorophyll biosynthesis and chloroplast development are two complex processes that are regulated by exogenous and endogenous factors. In this study, we identified OsDXR, a gene encoding a reductoisomerase that positively regulates chlorophyll biosynthesis and chloroplast development in rice. OsDXR knock-out lines displayed the albino phenotype and could not complete the whole life cycle process. OsDXR was highly expressed in rice leaves, and subcellular localization indicated that OsDXR is a chloroplast protein. Many genes involved in chlorophyll biosynthesis and chloroplast development were differentially expressed in the OsDXR knock-out lines compared to the wild type. Moreover, we found that the RNA editing efficiencies of ndhA-1019 and rpl2-1 were significantly reduced in the OsDXR knock-out lines. Furthermore, OsDXR interacted with the RNA editing factor OsMORF1 in a yeast two-hybrid screen and bimolecular fluorescence complementation assay. Finally, disruption of the plastidial 2-C-methyl-derythritol-4-phosphate pathway resulted in defects in chloroplast development and the RNA editing of chloroplast genes.
Photosynthesis occurs mainly in chloroplasts, whose development is regulated by proteins encoded by nuclear genes. Among them, pentapeptide repeat (PPR) proteins participate in organelle RNA editing. Although there are more than 450 members of the PPR protein family in rice, only a few affect RNA editing in rice chloroplasts. Gene editing technology has created new rice germplasm and mutants, which could be used for rice breeding and gene function study. This study evaluated the functions of OsPPR9 in chloroplast RNA editing in rice. The osppr9 mutants were obtained by CRISPR/Cas9, which showed yellowing leaves and a lethal phenotype, with suppressed expression of genes associated with chloroplast development and accumulation of photosynthetic-related proteins. In addition, loss of OsPPR9 protein function reduces the editing efficiency of rps8-C182, rpoC2-C4106, rps14-C80, and ndhB-C611 RNA editing sites, which affects chloroplast growth and development in rice. Our data showed that OsPPR9 is highly expressed in rice leaves and encodes a DYW-PPR protein localized in chloroplasts. Besides, the OsPPR9 protein was shown to interact with OsMORF2 and OsMORF9. Together, our findings provide insights into the role of the PPR protein in regulating chloroplast development in rice.
OsNPF3.1, a nitrate, abscisic acid and gibberellin transporter gene, is essential for rice tillering and nitrogen utilization efficiency
Low-affinity nitrate transporter genes have been identified in subfamilies 4–8 of the rice nitrate transporter 1 (NRT1)/peptide transporter family (NPF), but the OsNPF3 subfamily responsible for nitrate and phytohormone transport and rice growth and development remains unknown. In this study, we described OsNPF3.1 as an essential nitrate and phytohormone transporter gene for rice tillering and nitrogen utilization efficiency (NUtE). OsNPF3.1 possesses four major haplotypes of its promoter sequence in 517 cultivars, and its expression is positively associated with tiller number. Its expression was higher in the basal part, culm, and leaf blade than in other parts of the plant, and was strongly induced by nitrate, abscisic acid (ABA) and gibberellin 3 (GA3) in the root and shoot of rice. Electrophysiological experiments demonstrated that OsNPF3.1 is a pH-dependent low-affinity nitrate transporter, with rice protoplast uptake assays showing it to be an ABA and GA3 transporter. OsNPF3.1 overexpression significantly promoted ABA accumulation in the roots and GA accumulation in the basal part of the plant which inhibited axillary bud outgrowth and rice tillering, especially at high nitrate concentrations. The NUtE of OsNPF3.1-overexpressing plants was enhanced under low and medium nitrate concentrations, whereas the NUtE of OsNPF3.1 clustered regularly interspaced short palindromic repeats (CRISPR) plants was increased under high nitrate concentrations. The results indicate that OsNPF3.1 transports nitrate and phytohormones in different rice tissues under different nitrate concentrations. The altered OsNPF3.1 expression improves NUtE in the OsNPF3.1-overexpressing and CRISPR lines at low and high nitrate concentrations, respectively.
Artificial selection of the Green Revolution gene Semidwarf 1 is implicated in upland rice breeding
Semidwarf breeding has boosted crop production and is a well-known outcome from the first Green Revolution. The Green Revolution gene Semidwarf 1 (SD1), which modulates gibberellic acid (GA) biosynthesis, plays a principal role in determining rice plant height. Mutations in SD1 reduce rice plant height and promote lodging resistance and fertilizer tolerance to increase grain production. The plant height mediated by SD1 also favors grain yield under certain conditions. However, it is not yet known whether the function of SD1 in upland rice promotes adaptation and grain production. In this study, the plant height and grain yield of irrigated and upland rice were comparatively analyzed under paddy and dryland conditions. In response to dryland environments, rice requires a reduction in plant height to cope with water deficits. Upland rice accessions had greater plant heights than their irrigated counterparts under both paddy and dryland conditions, and appropriately reducing plant height could improve adaptability to dryland environments and maintain high grain yield formation. Moreover, upland rice cultivars with thicker stem diameters had stronger lodging resistance, which addresses the lodging problem. Knockout of SD1 in the upland rice cultivar IRAT104 reduced the plant height and grain yield, demonstrating that the adjustment of plant height mediated by SD1 could increase grain production in dryland fields. In addition, an SD1 genetic diversity analysis verified that haplotype variation causes phenotypic variation in plant height. During the breeding history of rice, SD1 allelic mutations were selected from landraces to improve the grain yield of irrigated rice cultivars, and this selection was accompanied by a reduction in plant height. Thus, five known mutant alleles were analyzed to verify that functional SD1 is required for upland rice production. All these results suggest that SD1 might have undergone artificial positive selection in upland rice, which provides further insights concerning greater plant height in upland rice breeding.
The auxin transporter OsAUX1 regulates tillering in rice (Oryza sativa)
Flower organ identity in rice is mainly determined by the A-, B-, C- and E-class genes, with the majority encoding MADS-box transcription factors. However, few studies have investigated how the expression of these floral organ identity genes is regulated during flower development. In this study, we identified a gene named SUPER WOMAN 2 (SPW2), which is necessary for spikelet/floret development in rice by participating in the regulation of the expression of pistil identity genes such as OsMADS3, OsMADS13, OsMADS58 and DL. In the spw2 mutant, ectopic stigma/ovary-like tissues were observed in the non-pistil organs, including sterile lemma, lemma, palea, lodicule, and stamen, suggesting that the identities of these organs were severely affected by mutations in SPW2. SPW2 was shown to encode a plant-specific EMF1-like protein that is involved in H3K27me3 modification as an important component of the PRC2 complex. Expression analysis showed that the SPW2 mutation led to the ectopic expression of OsMADS3, OsMADS13, OsMADS58, and DL in non-pistil organs of the spikelet. The ChIP-qPCR results showed significant reductions in the levels of H3K27me3 modification on the chromatin of these genes. Thus, we demonstrated that SPW2 can mediate the process of H3K27me3 modification of pistil-related genes to regulate their expression in non-pistil organs of spikelets in rice. The results of this study expand our understanding of the molecular mechanism by which SPW2 regulates floral organ identity genes through epigenetic regulation.