RNAi-directed downregulation of OsBADH2 results in aroma (2-acetyl-1-pyrroline) production in rice (Oryza sativa L.)

<p>Abstract</p> <p>Background</p> <p>Aromatic rice is popular worldwide because of its characteristic fragrance. Genetic studies and physical fine mapping reveal that a candidate gene (<it>fgr</it>/<it>OsBADH2</it>) homologous to <it>betaine aldehyde dehydrogenase </it>is responsible for aroma metabolism in fragrant rice varieties, but the direct evidence demonstrating the functions of <it>OsBADH2 </it>is lacking. To elucidate the physiological roles of <it>OsBADH2</it>, sequencing approach and RNA interference (RNAi) technique were employed to analyze allelic variation and functions of <it>OsBADH2 </it>gene in aroma production. Semi-quantitative, real-time reverse transcription-polymerase chain reaction (RT-PCR), as well as gas chromatography-mass spectrometry (GC-MS) were conducted to determine the expression levels of <it>OsBADH2 </it>and the fragrant compound in wild type and transgenic <it>OsBADH2</it>-RNAi repression lines, respectively.</p> <p>Results</p> <p>The results showed that multiple mutations identical to <it>fgr </it>allele occur in the 13 fragrant rice accessions across China; <it>OsBADH2 </it>is expressed constitutively, with less expression abundance in mature roots; the disrupted <it>OsBADH2 </it>by RNA interference leads to significantly increased 2-acetyl-1-pyrroline production.</p> <p>Conclusion</p> <p>We have found that the altered expression levels of <it>OsBADH2 </it>gene influence aroma accumulation, and the prevalent aromatic allele probably has a single evolutionary origin.</p

Cell Wall Matrix Polysaccharides Contribute to Salt–Alkali Tolerance in Rice

Salt–alkali stress threatens the resilience to variable environments and thus the grain yield of rice. However, how rice responds to salt–alkali stress at the molecular level is poorly understood. Here, we report isolation of a novel salt–alkali-tolerant rice (SATR) by screening more than 700 germplasm accessions. Using 93-11, a widely grown cultivar, as a control, we characterized SATR in response to strong salt–alkali stress (SSAS). SATR exhibited SSAS tolerance higher than 93-11, as indicated by a higher survival rate, associated with higher peroxidase activity and total soluble sugar content but lower malonaldehyde accumulation. A transcriptome study showed that cell wall biogenesis-related pathways were most significantly enriched in SATR relative to 93-11 upon SSAS. Furthermore, higher induction of gene expression in the cell wall matrix polysaccharide biosynthesis pathway, coupled with higher accumulations of hemicellulose and pectin as well as measurable physio-biochemical adaptive responses, may explain the strong SSAS tolerance in SATR. We mapped SSAS tolerance to five genomic regions in which 35 genes were candidates potentially governing SSAS tolerance. The 1,4-β-D-xylan synthase gene OsCSLD4 in hemicellulose biosynthesis pathway was investigated in details. The OsCSLD4 function-disrupted mutant displayed reduced SSAS tolerance, biomass and grain yield, whereas the OsCSLD4 overexpression lines exhibited increased SSAS tolerance. Collectively, this study not only reveals the potential role of cell wall matrix polysaccharides in mediating SSAS tolerance, but also highlights applicable value of OsCSLD4 and the large-scale screening system in developing SSAS-tolerant rice

Anti-allergic actions of a Chinese patent medicine, huoxiangzhengqi oral liquid, in RBL-2H3 cells and in mice

Context Huoxiangzhengqi oral liquid (HXZQ-OL), a traditional Chinese medicine formula, has antibacterial, anti-inflammation and gastrointestinal motility regulation effects. Objective The study investigates the anti-allergic activity and underlying mechanism of HXZQ-OL. Materials and methods IgE/Ag-mediated RBL-2H3 cells were used to evaluate the anti-allergic activity of HXZQ-OL (43.97, 439.7 and 4397 μg/mL) in vitro. The release of cytokines and eicosanoids were quantified using ELISA. RT-qPCR was used to measure the gene expression of cytokines. The level of intracellular Ca2+ was measured with Fluo 3/AM. Immunoblotting analysis was performed to investigate the mechanism of HXZQ-OL. In the passive cutaneous anaphylaxis (PCA), BALB/c mice (5 mice/group) were orally administrated with HXZQ-OL (263.8, 527.6 and 1055 mg/kg/d) or dexamethasone (5 mg/kg/d, positive control) for seven consecutive days. Results HXZQ-OL not only inhibited degranulation of mast cells (IC50, 123 μg/mL), but also inhibited the generation and secretion of IL-4 (IC50, 171.4 μg/mL), TNF-α (IC50, 88.4 μg/mL), LTC4 (IC50, 52.9 μg/mL) and PGD2 (IC50, 195.8 μg/mL). Moreover, HXZQ-OL suppressed the expression of IL-4 and TNF-α mRNA, as well as the phosphorylation of Fyn, Lyn and multiple downstream signalling proteins including MAPK and PI3K/NF-κB pathways. In addition, HXZQ-OL (527.5 mg/kg) attenuated the IgE-mediated PCA with 55% suppression of Evans blue exudation in mice. Conclusions HXZQ-OL attenuated the activation of mast cell and PCA. Therefore, HXZQ-OL might be used as an alternative treatment for allergic diseases

<i>OsMADS1</i> Regulates Grain Quality, Gene Expressions, and Regulatory Networks of Starch and Storage Protein Metabolisms in Rice

OsMADS1 plays a vital role in regulating floret development and grain shape, but whether it regulates rice grain quality still remains largely unknown. Therefore, we used comprehensive molecular genetics, plant biotechnology, and functional omics approaches, including phenotyping, mapping-by-sequencing, target gene seed-specific RNAi, transgenic experiments, and transcriptomic profiling to answer this biological and molecular question. Here, we report the characterization of the ‘Oat-like rice’ mutant, with poor grain quality, including chalky endosperms, abnormal morphology and loose arrangement of starch granules, and lower starch content but higher protein content in grains. The poor grain quality of Oat-like rice was found to be caused by the mutated OsMADS1Olr allele through mapping-by-sequencing analysis and transgenic experiments. OsMADS1 protein is highly expressed in florets and developing seeds. Both OsMADS1-eGFP and OsMADS1Olr-eGFP fusion proteins are localized in the nucleus. Moreover, seed-specific RNAi of OsMADS1 also caused decreased grain quality in transgenic lines, such as the Oat-like rice. Further transcriptomic profiling between Oat-like rice and Nipponbare grains revealed that OsMADS1 regulates gene expressions and regulatory networks of starch and storage protein metabolisms in rice grains, hereafter regulating rice quality. In conclusion, our results not only reveal the crucial role and preliminary mechanism of OsMADS1 in regulating rice grain quality but also highlight the application potentials of OsMADS1 and the target gene seed-specific RNAi system in improving rice grain quality by molecular breeding