Oral Administration of Germinated, Pigmented, Giant Embryo Rice (Oryza sativa L. cv. Keunnunjami) Extract Improves the Lipid and Glucose Metabolisms in High-Fat Diet-Fed Mice.

Obesity is a significant risk factor for chronic diseases. The effect of ethanol extract from germinated Keunnunjami, blackish-purple rice with a giant embryo, compare to ordinary brown rice, on the body weight and lipid and glucose metabolism in high-fat diet-fed mice was analyzed. Mice were fed with a high-fat diet-fed for 3 weeks and then orally administered with either distilled water (HF) or extract (0.25%, w/w) from brown, germinated brown, Keunnunjami, and germinated Keunnunjami rice for 4 weeks. Control mice were fed with a normal diet and orally administered with distilled water. The HF group showed markedly higher body weight and triglyceride, cholesterol, fatty acid, glucose, and insulin levels than the control group. However, the oral administration of rice extracts ameliorated this high-fat diet-induced obesity, hyperlipidemia, and hypoglycemia through the modulation of adipokine production, lipogenic and glucose-regulating enzyme activities, and mRNA expression of genes associated with lipid and glucose metabolism. The germinated Keunnunjami extract exhibited greater hypolipidemic, hypoglycemic, and body weight-lowering effects than the other rice extracts. The results demonstrated that germination could further enhance the physiological properties of rice and that germinated Keunnunjami extract has a strong therapeutic potential against high-fat diet-induced obesity, hyperlipidemia, and hyperglycemia.

CHR721, interacting with OsRPA1a, is essential for both male and female reproductive development in rice.

KEY MESSAGE: CHR721 functions as a chromatin remodeler and interacts with a known single-stranded binding protein, OsRPA1a, to regulate both male and female reproductive development in rice. Reproductive development and fertility are important for seed production in rice. Here, we identified a sterile rice mutant, chr721, that exhibited defects in both male and female reproductive development. Approximately 5% of the observed defects in chr721, such as asynchronous dyad division, occurred during anaphase II of meiosis. During the mitotic stage, approximately 80% of uninucleate microspores failed to develop into tricellular pollen, leading to abnormal development. In addition, defects in megaspore development were detected after functional megaspore formation. CHR721, which encodes a nuclear protein belonging to the SNF2 subfamily SMARCAL1, was identified by map-based cloning. CHR721 was expressed in various tissues, especially in spikelets. CHR721 was found to interact with replication protein A (OsRPA1a), which is involved in DNA repair. The expressions of genes involved in DNA repair and cell-cycle checkpoints were consistently upregulated in chr721. Although numerous genes involved in male and female development have been identified, the mode of participation of chromatin-remodeling factors in reproductive development is still not well understood. Our results suggest that CHR721, a novel gene cloned from rice, plays a vital role in both male and female reproductive development.

Changes in mineral elements and starch quality of grains during the improvement of japonica rice cultivars.

BACKGROUND: The improvement of rice cultivars plays an important role in yield increase. However, little is known about the changes in starch quality and mineral elements during the improvement of rice cultivars. This study was conducted to investigate the changes in starch quality and mineral elements in japonica rice cultivars. RESULTS: Twelve typical rice cultivars, applied in the production in Jiangsu province during the last 60 years, were grown in the paddy fields. These cultivars were classified into six types according to their application times, plant types and genotypes. The nitrogen (N), phosphorus (P) and, and potassium (K) were mainly distributed in endosperm, bran and bran, respectively. Secondary and micromineral nutrients were distributed throughout grains. With the improvement of cultivars, total N contents gradually decreased, while total P, K and magnesium contents increased in grains. Total copper and zinc contents in type 80'S in grains were highest. The improvement of cultivars enhanced palatability (better gelatinisation enthalpy and amylose content), taste (better protein content) and protein quality (better protein components and essential amino acids). Correlation analysis indicated the close relationship between mineral elements and starch quality. CONCLUSION: The mineral elements and starch quality of grains during the improvement of japonica rice cultivars are improved. © 2017 Society of Chemical Industry.

Acyl-CoA-Binding Proteins (ACBPs) in Plant Development.

Acyl-CoA-binding proteins (ACBPs) play a pivotal role in fatty acid metabolism because they can transport medium- and long-chain acyl-CoA esters. In eukaryotic cells, ACBPs are involved in intracellular trafficking of acyl-CoA esters and formation of a cytosolic acyl-CoA pool. In addition to these ubiquitous functions, more specific non-redundant roles of plant ACBP subclasses are implicated by the existence of multigene families with variable molecular masses, ligand specificities, functional domains (e.g. protein-protein interaction domains), subcellular locations and gene expression patterns. In this chapter, recent progress in the characterization of ACBPs from the model dicot plant, Arabidopsis thaliana, and the model monocot, Oryza sativa, and their emerging roles in plant growth and development are discussed. The functional significance of respective members of the plant ACBP families in various developmental and physiological processes such as seed development and germination, stem cuticle formation, pollen development, leaf senescence, peroxisomal fatty acid ß-oxidation and phloem-mediated lipid transport is highlighted.

The role of OsMSH4 in male and female gamete development in rice meiosis.

Meiosis is essential for gametogenesis in sexual reproduction in rice (Oryza sativa L.). We identified a MutS-homolog (MSH) family gene OsMSH4 in a trisomic plant. Cytological analysis showed that developments of both pollen and embryo sacs in an Osmsh4 mutant were blocked due to defective chromosome pairing. Compared with the wild type, the Osmsh4 mutant displayed a significant ~21.9% reduction in chiasma frequency, which followed a Poisson distribution, suggesting that class I crossover formation in the mutant was impaired. Temporal and spatial expression pattern analyses showed that OsMSH4 was preferentially expressed in meiocytes during their meiosis, indicating a critical role in gametogenesis. Subcellular localization showed that OsMSH4-green fluorescent protein was predominantly located in the nucleus. OsMSH4 could interact with another MSH member (OsMSH5) through the N-terminus and C-terminus, respectively. Direct physical interaction between OsMSH5, OsRPA1a, OsRPA2b, OsRPA1c, and OsRPA2c was identified by yeast two-hybrid assays and further validated by pull-down assays. Our results supported the conclusion that the OsMSH4/5 heterodimer plays a key role in regulation of crossover formation during rice meiosis by interaction with the RPA complex.

Effects of silicon on seed setting rate of rice intersubspecific hybrids.

The present study found semi-sterility in rice intersubspecific hybrids of 'Taichung 65' x 'Guangluai 4' and 'Ludao' x 'Qiuguang'. Embryo sac fertility was evaluated using the overall staining transparent method. The results showed that the embryo sac contained a normal egg cell, normal synergid cells, polar nuclei cells, and antipodal cells, indicating that semi-sterility was caused mainly by pollen semi-sterility. In the pot experiment, the effects of silicon on the seed-setting rate of the two intersubspecific hybrids were examined. The results showed that the rate of anther dehiscence, number of pollen per stigma of Fl plants, potential of pollen grain germination, and fertility of the spikelet were significantly improved by the utilization of silicon fertilizer.

Amyloplast-localized SUBSTANDARD STARCH GRAIN4 protein influences the size of starch grains in rice endosperm.

Starch is a biologically and commercially important polymer of glucose and is synthesized to form starch grains (SGs) inside amyloplasts. Cereal endosperm accumulates starch to levels that are more than 90% of the total weight, and most of the intracellular space is occupied by SGs. The size of SGs differs depending on the plant species and is one of the most important factors for industrial applications of starch. However, the molecular machinery that regulates the size of SGs is unknown. In this study, we report a novel rice (Oryza sativa) mutant called substandard starch grain4 (ssg4) that develops enlarged SGs in the endosperm. Enlargement of SGs in ssg4 was also observed in other starch-accumulating tissues such as pollen grains, root caps, and young pericarps. The SSG4 gene was identified by map-based cloning. SSG4 encodes a protein that contains 2,135 amino acid residues and an amino-terminal amyloplast-targeted sequence. SSG4 contains a domain of unknown function490 that is conserved from bacteria to higher plants. Domain of unknown function490-containing proteins with lengths greater than 2,000 amino acid residues are predominant in photosynthetic organisms such as cyanobacteria and higher plants but are minor in proteobacteria. The results of this study suggest that SSG4 is a novel protein that influences the size of SGs. SSG4 will be a useful molecular tool for future starch breeding and biotechnology.

Regeneration of plantlets from mature embryo calli of Western Ghats land race cultivar of rice, Oryza sativa L.

The Malnad region located in the Western Ghats of Karnataka is known for the cultivation of indigenous rain fed land race cultivar of rice. The present study was to investigate the callogenic and caulogenic potentialities of the two indigenous rice cultivar namely Karimundaga and Kanadatumba using dehusked mature embryo explants. For callus and shoot bud differentiation, the explants were cultured on Murashige and Skoog (MS) medium supplemented with 2,4-D (1-3 mg/L), IAA (1-2 mg/L), Kn (1-4 mg/L) and BAP (1-4 mg/L). The morphogenic potentialities of the two rice cultivar differed in texture of callus. In both the cultivar callogenic frequency was optimized at 1 mg/L 2,4-D concentration, it was 94% in Karimundaga and 58% in Kanadatumba. Supplementation of IAA either alone (1-2 mg/L) or in combination with Kn or BAP at 1 to 4 mg/L concentration of each induces shoot bud differentiation from the calli. In the cultivar Karimundaga caulogenic frequency was highest (10.60 +/- 2.55) at 1.0 mg/L IAA and 4.0 mg/L BAP concentration. While in the cultivar Kanadatumba highest number of shoot buds (7.90 +/- 2.69) was differentiated at 1.0 mg/L IAA and 4.0 mg/L Kn concentration. The calli derived regenerants were successfully acclimatized in the greenhouse and agro-morphological variations were evaluated. The growth characteristics and yield related parameters exhibited by in vitro plants were lower than the in vivo plants.

Replication protein A (RPA1a) is required for meiotic and somatic DNA repair but is dispensable for DNA replication and homologous recombination in rice.

Replication protein A (RPA), a highly conserved single-stranded DNA-binding protein in eukaryotes, is a stable complex comprising three subunits termed RPA1, RPA2, and RPA3. RPA is required for multiple processes in DNA metabolism such as replication, repair, and homologous recombination in yeast (Saccharomyces cerevisiae) and human. Most eukaryotic organisms, including fungi, insects, and vertebrates, have only a single RPA gene that encodes each RPA subunit. Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa), however, possess multiple copies of an RPA gene. Rice has three paralogs each of RPA1 and RPA2, and one for RPA3. Previous studies have established their biochemical interactions in vitro and in vivo, but little is known about their exact function in rice. We examined the function of OsRPA1a in rice using a T-DNA insertional mutant. The osrpa1a mutants had a normal phenotype during vegetative growth but were sterile at the reproductive stage. Cytological examination confirmed that no embryo sac formed in female meiocytes and that abnormal chromosomal fragmentation occurred in male meiocytes after anaphase I. Compared with wild type, the osrpa1a mutant showed no visible defects in mitosis and chromosome pairing and synapsis during meiosis. In addition, the osrpa1a mutant was hypersensitive to ultraviolet-C irradiation and the DNA-damaging agents mitomycin C and methyl methanesulfonate. Thus, our data suggest that OsRPA1a plays an essential role in DNA repair but may not participate in, or at least is dispensable for, DNA replication and homologous recombination in rice.

Efficient in vitro plant regeneration via leaf base segments of indica rice (Oryza sativa L.).

A reliable and reproducible protocol has been developed for high frequency plant regeneration from 4-5 mm long leaf base segments of 4 days old in vitro germinated seedlings of indica rice (Oryza sativa) cultivar Rasi. The effect of age of seedlings, position of leaf base segments and optimum concentration of 2,4-D on callus induction frequency was investigated with a future aim to use leaf bases for biolistic and Agrobacterium-mediated transformation experiments. Friable, nodular and white to pale yellow embryogenic callus cultures (206 mg fresh weight /explant) were obtained from the first basal segments of rice seedlings on Linsmaier and Skoog (LS) medium supplemented with 2,4-D (11.3 microM) and 3.0 microM thiamine-HCL. Plant regeneration was achieved after the transfer of 54 days old embryogenic callus cultures to Murashige and Skoog (MS) medium supplemented with BAP (2.2 microM) and NAA (0.27 microM). In vitro regenerated plants with multiple shoots and roots transferred to sterile soil in growth chamber and maintained in greenhouse exhibited normal growth and were phenotypically similar to plants germinated from seeds.

Effect of 2,4-d, hydric stress and light on indica rice (Oryza sativa) somatic embryogenesis.

With the purpose of increasing the embryogenesis regeneration process in vitroplants obtained from somatic embryos of the indica rice variety CR-5272 (Oryza sativa L.), two independent experiments were performed. The first experiment consisted in the effect of combination of three concentrations of the gelling agent Phytagel (1.8. 2.4. and 3 gL(-1)) and four 2,4-D concentrations (2.26, 4.52. 6.78, and 9.05 microM) on the induction and subsequent regeneration of embryogenic calli. On the second experiment, the pre-regeneration phase was modified: calli were subjected to darkness or diffuse light conditions for one, two, and three weeks. In embryogenesis induction, 35% calligenesis was obtained using the MS culture medium supplemented with 6.78 microM of 2.4-D and 2.4 gL(-1) Phytagel, whereas on the control treatment (MS medium supplemented with 9.05 microM of 2.4-D and 3 gL(-1) Phytagel ) 24% calligenesis was obtained. In addition, regeneration percentages were improved (22% and 16% for calli induced with the above treatments, respectively). Furthermore, in light exposure experiments, the best result was obtained by exposing the embryogenic calli to darkness for one week in pre-regeneration, followed by direct light exposure during the regeneration phase.

A new 9-lipoxygenase cDNA from developing rice seeds.

We isolated a novel C9 position specific lipoxygenase (r9-LOX1) cDNA from developing rice seeds. The enzymatic features of r9-LOX1 resembled those of rice LOX-L3 known to be contained in rice germ and to have C9-specific LOX activity. However, the expression level of the r9-LOX1 gene was higher in imbibed seeds rather than developing seeds. A homology search against the rice nucleotide database revealed the r9-LOX1 gene to be on rice chromosome 3 (accession number AC093017). The restriction enzyme map of the reported genomic sequence agreed with the result of the Southern blot analysis for the r9-LOX1. The enzyme could be useful for in vitro synthesis of 9,10-ketol-octadecadienoic acid.

Brassinolide improves embryogenic tissue initiation in conifers and rice.

Somatic embryogenesis (SE), the most promising technology for the large-scale production of high-value coniferous trees from advanced breeding and genetic engineering programs, is expected to play an important role in increasing productivity, sustainability, and the uniformity of future U.S. forests. To be successful for commercial use, SE technology must work with a variety of genetically diverse trees. Initiation in loblolly pine ( Pinus taeda L.), our main focus species, is often recalcitrant for desirable genotypes. Initiation percentages of loblolly pine, Douglas-fir [ Pseudotsuga menziesii (Mirb.) Franco], and Norway spruce ( Picea abies L., Karst.) were improved through the use of brassinolide. Brassinosteroids, which include brassinolide, are a relatively new group of natural plant growth regulators that are found in many plant species. They have been shown to have diverse, tissue-specific, and species-specific effects, including the stimulation of cell elongation and ethylene production and increasing resistance to abiotic stress. In our media, brassinolide was effective at concentrations ranging from 0.005-0.25 micro M. Using control medium (no brassinolide) and brassinolide-supplemented (0.1 micro M) medium, we achieved improved initiation percentages in loblolly pine, Douglas-fir, Norway spruce, and rice-15.0% to 30.1%, 16.1% to 36.3%, 34.6% to 47.4%, and 10%, respectively. Brassinolide increased the weight of loblolly pine embryogenic tissue by 66% and stimulated initiation in the more recalcitrant families of loblolly pine and Douglas-fir, thus compensating somewhat for genotypic differences in initiation. Initiation percentages in loblolly pine were improved through the combination of modified 1/2-P6 salts, 50 mg/l activated carbon (AC), adjusted levels of Cu and Zn (to compensate for adsorption by AC), 1.5% maltose, 2% myo-inositol (to raise the osmotic level, partially simulating the megagametophyte environment), 500 mg/l casamino acids, 450 mg/l glutamine, 2 mg/l alpha-naphthaleneacetic acid, 0.63 mg/l 6-benzylaminopurine, 0.61 mg/l kinetin, 3.4 mg/l silver nitrate, 10 micro M cGMP, 0.1 micro M brassinolide, and 2 g/l Gelrite. Across 12 open-pollinated families of loblolly pine, initiation percentages ranged from 2.5% to 50.7%, averaging 22.5%.

Novel gene encoding a Ca2+-binding protein and under hexokinase-dependent sugar regulation.

A cDNA encoding a predicted 15-kDa protein was earlier isolated from sugar-induced genes in rice embryos (Oryza sativa L.) by cDNA microarray analysis. Here we report that this cDNA encodes a novel Ca2+-binding protein, named OsSUR1 (for Oryza sativa sugar-up-regulated-1). The recombinant OsSUR1 protein expressed in Escherichia coli had 45Ca2+-binding activity. Northern analysis showed that the OsSUR1 gene was expressed mainly in the internodes of mature plants and in embryos at an early stage of germination. Expression of the OsSUR1 gene was induced by sugars that could serve as substrates of hexokinase, but expression was not repressed by Ca2+ signaling inhibitors, calmodulin antagonists and inhibitors of protein kinase or protein phosphatase. These results suggested that Os-SUR1 gene expression was stimulated by a hexokinase-dependent pathway not mediated by Ca2+.