Among three genes for cytosolic glutamine synthetase (OsGS1;1, OsGS1;2 and OsGS1;3) in rice (Oryza sativa L.) plants, the OsGS1;2 gene is known to be mainly expressed in surface cells of roots, but its function was not clearly understood. We characterized knock-out mutants caused by the insertion of an endogenous retrotransposon Tos17 into exon 2 of OsGS1;2. Homozygously inserted mutants showed severe reduction in active tiller number and hence panicle number at harvest. Other yield components, such as spikelet number per panicle, 1,000-spikelet weight and proportion of well ripened grains, were nearly identical between the mutants and wild-type plants. When the contents of free amino acids in roots were compared between the mutants and the wild type, there were marked reductions in contents of glutamine, glutamate, asparagine and aspartate, but a remarkable increase in free ammonium ions in the mutants. Concentrations of amino acids and ammonium ions in xylem sap behaved in a similar fashion. Re-introduction of OsGS1;2 cDNA under the control of its own promoter into the knock-out mutants successfully restored yield components to wild-type levels as well as ammonium concentration in xylem sap. The results indicate that GS1;2 is important in the primary assimilation of ammonium ions taken up by rice roots, with GS1;1 in the roots unable to compensate for GS1;2 functions.

;To characterize ammonium transport pathways in rice, two cDNAs with high homology to MEP/AMT2-type ammonium transporters, OsAMT2;1 and OsAMT3;1, were isolated. Expression of OsAMT2;1 in an ammoniumuptake-defective yeast mutant showed that this gene encodes functional ammonium transporters. OsAMT2;1 was constitutively expressed in both roots and shoots irrespective of the supply of inorganic nitrogen to the medium, whereas OsAMT3;1 expression was relatively weak. A database search with the amino acid sequence of OsAMT2;1 showed that there are 10 putative OsAMT genes in rice, i.e. three each for OsAMT1, OsAMT2 and OsAMT3, respectively, and one for OsAMT4.

Immunocytological studies in this laboratory have suggested that NADH-dependent glutamate synthase (NADH-GOGAT; EC 1.4.1.14) in developing organs of rice (Oryza sativa L. cv. Sasanishiki) is involved in the utilization of glutamine remobilized from senescing organs through the phloem. Because most of the indica cultivars contained less NADH-GOGAT in their sink organs than japonica cultivars, over-expression of NADH-GOGAT gene from japonica rice was investigated using Kasalath, an indica cultivar. Several T0 transgenic Kasalath lines over-producing NADH-GOGAT under the control of a NADH-GOGAT promoter of Sasanishiki, a japonica rice, showed an increase in grain weight (80% as a maximum), indicating that NADH-GOGAT is indeed a key step for nitrogen utilization and grain filling in rice. A genetic approach using 98 backcross-inbred lines (BC(1)F(6)) developed between Nipponbare (a japonica rice) and Kasalath were employed to detect putative quantitative trait loci (QTLs) associated with the contents of cytosolic glutamine synthetase (GS1; EC 6.3.1.2), which is probably involved in the export of nitrogen from senescing organs and those of NADH-GOGAT. Immunoblotting analyses showed transgressive segregations toward lower or greater contents of these enzyme proteins in these BC(1)F(6). Seven chromosomal QTL regions were detected for GS1 protein content and six for NADH-GOGAT. Some of these QTLs were located in QTL regions for various biochemical and agronomic traits affected by nitrogen recycling. The relationships between the genetic variability of complex agronomic traits and traits for these two enzymes are discussed.

Background Rikkunshito, a standardized Japanese herbal medicine, is thought to accelerate gastric emptying and relieve dyspepsia, although no large‐scale, randomized, placebo‐controlled trials of rikkunshito have been conducted. This study aimed to determine the efficacy and safety of rikkunshito for treating functional dyspepsia (FD). Methods FD patients received 2.5 g rikkunshito or placebo three times a day for 8 weeks in this multicenter, randomized, placebo‐controlled, parallel‐group trial. The primary end point was the proportion of responders at 8 weeks after starting test drug, determined by global patient assessment (GPA). The improvement in four major dyspepsia symptoms severity scale was also evaluated. In addition, plasma ghrelin levels were investigated before and after treatment. Key Results Two hundred forty‐seven patients were randomly assigned. In the eighth week, the rikkunshito group had more GPA responders (33.6%) than the placebo (23.8%), although this did not reach statistical significance (p = 0.09). Epigastric pain was significantly improved (p = 0.04) and postprandial fullness tended to improve (p = 0.06) in the rikkunshito group at week 8. Rikkunshito was relatively more effective among Helicobacter pylori‐infected participants (rikkunshito: 40.0% vs placebo: 20.5%, p = 0.07), and seemed less effective among H. pylori‐uninfected participants (rikkunshito: 29.3% vs placebo: 25.6%, p = 0.72). Among H. pylori‐positive individuals, acyl ghrelin levels were improved just in rikkunshito group. There were no severe adverse events in both groups. Conclusions & Inferences Administration of rikkunshito for 8 weeks reduced dyspepsia, particularly symptoms of epigastric pain and postprandial fullness. (UMIN Clinical Trials Registry, Number UMIN000003954).

Rice (Oryza sativa L.) plants with substantially increased Rubisco content were obtained by Agrobacteriummediated transformation with the rice rbcS sense gene under the control of the rice rbcS promoter. The primary transformants were screened for the ratio of Rubisco to leaf-N content, and the transformants with 4120% wild-type levels of Rubisco were selected. In the progeny of the selected lines of the transformants, the mRNA levels of one member of the rbcS gene family were increased from 3.9-to 6.2-fold, whereas those of other members of the rbcS gene family were unchanged. The total levels of rbcS mRNA were increased from 2.1-to 2.8-fold. The levels of rbcL mRNA were increased from 1.2-to 1.9-fold. Rubisco protein content was significantly increased by 30% on a leaf area basis. The ratio of Rubisco-N to leaf-N was also increased by 10-20%, irrespective of N treatment. The specific activity of Rubisco per unit of enzyme protein was not different. However, light-saturated photosynthesis was not enhanced even when the rate was measured at low [CO 2 ] where Rubisco becomes limiting for photosynthesis. Some lines showed lower photosynthesis at high [CO 2 ] (460 Pa). We conclude that introduction of additional sense rbcS leads to overexpression of rbcS and that this overexpression slightly up-regulates the gene expression of rbcL at the transcript level and enhances the amount of Rubisco holoenzyme. However, overproduction of Rubisco protein does not improve photosynthesis.