Characterizing nicotianamine aminotransferase: Improving its assay system and details of the regulation of its activity by Fe nutrition status

Under iron deficient conditions, graminaceous plants secrete mugineic acid family phytosiderophores (MAs) from their roots to dissolve sparingly soluble iron compounds in the rhizosphere, and take up iron in the form of an Fe³⁺-MAs complex (Takagi 1976). A good correlation has been reported between the tolerance of Fe-deficiency and the amount of secreted MAs (Takagi 1993). Therefore, by using the genes involved in MAs biosynthesis, molecular breeding might produce transgenic plants tolerant to Fe-deficiency with a high level of MAs secretion. The biosynthetic pathway of MAs from L-methionine has been clarified (Fig. 1) and the enzymes participating in this process are now being investigated to isolate the genes responsible. Nicotianamine aminotransferase (NAAT) catalyzes the amino group transfer between nicotianamine (NA) and 2-oxoglutaric acid (Fig. 1). In order to purify NAAT, enzyme assay methods for NAAT have been developed and modified (Shojima et al. 1990; Ohata et al. 1993; Kanazawa et al. 1994). Some characteristics of NAAT have been reported using these enzyme assay methods (Kanazawa et al. 1994, 1995). Here, we further investigate some characteristics of this enzyme to improve the enzyme assay method, namely 1) the effect of K⁺ and Mg²⁺ on NAAT activity in vitro, and 2) the direct influence of MAs, Fe³⁺, and Fe²⁺ on NAAT activity. In addition, based on these results, the induction of enzyme activity by Fe-deficiency and suppression of the activity by Fe-resupply was investigated, by applying the new enzyme assay method.     

A rapid and easy method of detecting levels of gene expression in intact tobacco plants using an Agrobacterium-mediated transient assay: Toward development of nutritional diagnosis based on gene expression

For the use of gene expression for nutritional diagnosis, a rapid and easy method of detecting levels of gene expression was proposed. Agrobacterium tumefaciens carrying the GUS reporter gene was injected into intact tobacco leaves with a plastic syringe, and transient GUS expression was determined. GUS activities were reproducible in young leaves.     

Time course study of aluminum-induced callose formation in barley roots as observed by digital microscopy and low-vacuum scanning electron microscopy

To clarify the mechanism(s) involved in the short-term inhibition of root elongation by AI, we monitored the morphological changes of barley roots by digital microscopy. Within 30 min after exposure to 37 µM AI, the surface of the root epidermis in the region of a distance of 1.5 mm from the root tip became rough and began to show signs of damage. After 38 min, callose was rapidly excreted from the junction between the root cap and the root epidermis, and formed a spherical lump approximately 60 µm in diameter. The fine structure of the callose deposits on the root surface was analyzed by low-vacuum scanning electron microscopy. After 50 min, there was a significant increase in the callose contents in the distal 0.6 mm part. At the same time, root elongation stopped completely. Fluorescence staining indicated that callose was localized on the surface of the cell elongation area (the elongation zone of primary roots and root hairs), but not on the surface of the meristem. The root growth reduction associated with AI treatment may be due to the use of sugar substrates for callose formation instead of cellulose formation.     

Turnover time of microbial biomass carbon in Japanese upland soils with different textures

We investigated the turnover time of microbial biomass-C in Japanese upland soils with various textures and examined the soil physicochemical properties influencing their turnover time. Samples from five different soil types (sand-dune regosol, light-colored Andosol, humic Andosol, brown forest soil, and dark red soil) were taken from upland concrete-frame plots in the experimental field of Chiba University. Each soil amended with [U -¹³C] glucose was incubated for 80 d at 25°C. Microbial biomass-C and -¹³C in soil were periodically determined by the fumigation-extraction method. The longest turnover time of microbial biomass-C was observed in the dark red soil (215 d) followed by the humic Andosol (134 d), brown forest soil (97 d), and light-colored Andosol (83 d) and the shortest in the sand-dune regosol (45 d). The turnover time of microbial biomass-C was significantly correlated with the value of soil clay (R: 0.917*), CEC (R: 0.921*), and macroaggregate (R: 0.907*) contents, but not with the total-C content. The amount of microbial biomass-C showed a close correlation with the turnover time of microbial biomass-C, suggesting that the turnover time of microbial biomass-C is an important factor influencing the accumulation of microbial biomass-C in soil.     

Effect of free-air CO2 enrichment on the storage of carbohydrate fixed at different stages in rice (Oryza sativa L.)

Increasing global atmospheric CO₂ concentrations are expected to influence crop production. To investigate the effect on rice (Oryza sativa L.), plants were grown under ambient CO₂ (AMB) or free-air CO₂-enrichment (FACE) at CO₂ concentrations ranged from 275 to 365 μmol mol⁻¹ above AMB. We supplied ¹³CO₂ to the plants at different growth stages so we could examine the contribution of carbohydrate stored during the vegetative stage or newly fixed carbohydrate produced during the grain-filling stage to ear weight at grain maturity. In plants supplied with ¹³C at the panicle-initiation or pre-heading stages, plants grown under FACE had more starch in the stems at heading, but there was no difference in stem ¹³C content. Furthermore, there were no differences between treatments in whole-plant ¹³C contents at heading and grain maturity. In contrast, plants supplied with ¹³C at the grain-filling stage had more ¹³C in the whole plant and the ears at grain maturity under FACE than under AMB, indicating that the increased amount of photosynthate produced at the grain-filling stage under CO₂ enrichment might be effectively stored in the grains. Furthermore, regardless of when the ¹³C was supplied, plants had more ¹³C in starch in the ears at grain maturity under FACE than under AMB. Therefore, CO₂ enrichment appears to promote partitioning of photosynthate produced during both vegetative and grain-filling stages to the grains.     

Increase in Iron and Zinc Concentrations in Rice Grains Via the Introduction of Barley Genes Involved in Phytosiderophore Synthesis

Increasing the iron (Fe) and zinc (Zn) concentrations of staple foods, such as rice, could solve Fe and Zn deficiencies, which are two of the most serious nutritional problems affecting humans. Mugineic acid family phytosiderophores (MAs) play a very important role in the uptake of Fe from the soil and Fe transport within the plant in graminaceous plants. To explore the possibility of MAs increasing the Fe concentration in grains, we cultivated three transgenic rice lines possessing barley genome fragments containing genes for MAs synthesis (i.e., HvNAS1, HvNAS1, and HvNAAT-A and HvNAAT-B or IDS3) in a paddy field with Andosol soils. Polished rice seeds with IDS3 inserts had up to 1.40 and 1.35 times higher Fe and Zn concentrations, respectively, compared to non-transgenic rice seeds. Enhanced MAs production due to the introduced barley genes is suggested to be effective for increasing Fe and Zn concentrations in rice grains.     

Effect of sodium acetate on the adhesion to porcine gastric mucin in a Lactococcus lactis strain grown on fructose

The association of lactic acid bacteria with mucosal surfaces plays important roles in the beneficial effects of these bacteria on human health, such as colonization of the gastrointestinal tract for pathogen antagonism. Previously, we found that the adhesion of Lactococcus lactis 7‐1 to porcine gastric mucin was higher with fructose than with lactose, galactose or xylose as the carbon source. In this study, we examined the effect of growth conditions on the adhesion of strain 7‐1 grown on fructose. Medium components affect the adhesion: the adhesion of strain 7‐1 grown with sodium acetate was higher than that without it. The enhancement of adhesion by sodium acetate was not observed under aerobic conditions. Cellular properties grown with or without sodium acetate were characterized: strain 7‐1 grown with sodium acetate had similar sugar contents, and different fatty acid composition to those grown without it. Strain 7‐1 grown with sodium acetate showed significantly lower cell yield and significantly higher hydrophobicity than those grown without it, which is associated with higher adhesion. Fructose and sodium acetate are frequently used in the food industry; this study may reveal a simple way to enhance the adhesion of lactic acid bacteria by growing them with these substances.     

Development of furnished cages re‐using conventional cages for laying hens: Behaviour,physical condition and productivity

Furnished cages for laying hens have advantages in allowing normal behaviors and maintaining productivity. As the cost of introduction is a barrier for farms, we developed furnished cages that re‐use conventional cages. To determine the minimum and functional cage design, we compared six designs, combinations of two floor designs (artificial turf or wire cage floor) and three screening designs in the integrated area (no screening, one entrance side or four sides). In total, 144 hens were used, and we measured behavior, physical condition and productivity. Comparing the floors, the percentages of hens performing dust‐bathing and laying eggs in the integrated area were higher in cages with turf than wire floor (P < 0.05 for both). Comparing the screening, dust‐bathing, litter‐exploring and active behavior tended to be more frequent in cages with the integrated area screened on one side than four sides. Feather damage was lower in cages with the integrated area screened on one side than with no screening (P < 0.05). These results suggest that the cage design with an integrated area with artificial turf floor, screened on one side, was effective for furnished cages that re‐use conventional cages.     

不同地下水位胡杨蒸腾速率与叶水势的变化分析

本文研究了不同地下水位胡杨蒸腾速率、叶水势及蒸腾速率的季节变化等水分生理特性,结果表明:地下水位影响胡杨蒸腾作用,同时土壤水分状况也影响蒸腾速率,当地下水位在2.28~2.43 m时,胡杨蒸腾速率变化幅度大,蒸腾速率的日最大值为7.50 mmol.m-2s-1,日均值为4.73 mmol.m-2s-1,对应叶水势的日最低值为-3.55(Mpa)和日均值为-1.94(Mpa);地下水位2.91~3.33 m时,叶水势的日最低值为-2.61(Mpa)和日均值为-1.62(Mpa)。胡杨蒸腾速率季节变化在7月份达到最大值,靠近河岸胡杨蒸腾速率变化较远离河岸胡杨大,表明地下水位低时,胡杨保持较高的水势就能从环境中吸收到水分来满足其正常生长。     

Photosynthetic induction responses to variable light under field conditions in three species grown in the gap and understory of a Fagus crenata forest

Photosynthetic induction responses to abrupt increases in photon flux density (PFD) to 800 and 1500 &mgr;mol m(-2) s(-1) from either darkness or 100 &mgr;mol m(-2) s(-1) were examined in situ in leaves of Fagus crenata Blume, Daphniphyllum humile Maxim., and Acer rufinerve Siebold & Zucc. growing in a gap and the understory of an F. crenata forest. Among the species studied, F. crenata exhibited the highest assimilation rate (A(100)), stomatal conductance (g(s100)) at the background PFD of 100 &mgr;mol m(-2) s(-1), and A(100)/A(max) (A(max) = maximum assimilation rate), in both the gap and the understory. Time required for full induction depended on both background PFD and maximum PFD. The induction period was 2-4-fold shorter at a background PFD of 100 &mgr;mol m(-2) s(-1) than in darkness. For the three understory species, time required to full induction was 2-3-fold longer when irradiance was increased from darkness to 800 &mgr;mol m(-2) s(-1) than when irradiance was increased from darkness to 1500 &mgr;mol m(-2) s(-1). Acer rufinerve showed higher initial stomatal conductance (g(s0)) and a shorter induction period in the understory than in the gap. Fagus crenata exhibited a similar g(s0) and induction period in both habitats. Daphniphyllum humile demonstrated lower g(s0) and a longer induction period in the understory than in the gap. These findings indicate that initial stomatal conductance is closely correlated with the photosynthetic induction response. We conclude that the photosynthetic induction response is affected by the light conditions experienced by plants before the sudden increase in irradiance and by the extent of the increase in irradiance.