Natsuko I. Kobayashi

Leaf senescence in rice due to magnesium deficiency mediated defect in transpiration rate before sugar accumulation and chlorosis.

Magnesium (Mg) is an essential macronutrient supporting various functions, including photosynthesis. However, the specific physiological responses to Mg deficiency remain elusive. In this study, 2-week-old rice seedlings (Oryza sativa. cv. Nipponbare) with three expanded leaves (L2-L4) were transferred to Mg-free nutrient solution for 8 days. In the absence of Mg, on day 8, L5 and L6 were completely developed, while L7 just emerged. We also studied several mineral deficiencies to identify specific responses to Mg deficiency. Each leaf was analyzed in terms of chlorophyll, starch, anthocyanin and carbohydrate metabolites, and only absence of Mg was found to cause irreversible senescence of L5. Resupply of Mg at various time points confirmed that the borderline of L5 death was between days 6 and 7 of Mg deficiency treatment. Decrease in chlorophyll concentration and starch accumulation occurred simultaneously in L5 and L6 blades on day 8. However, nutrient transport drastically decreased in L5 as early as day 6. These data suggest that the predominant response to Mg deficiency is a defect in transpiration flow. Furthermore, changes in myo-inositol and citrate concentrations were detected only in L5 when transpiration decreased, suggesting that they may constitute new biological markers of Mg deficiency.

Estimation of radioactive 137-cesium transportation by litterfall, stemflow and throughfall in the forests of Fukushima.

Since the Fukushima Dai-ichi Nuclear Power Plant accident in March 2011, large areas of the forests around Fukushima have become highly contaminated by radioactive nuclides. To predict the future dynamics of radioactive cesium ((137)Cs) in the forest catchment, it is important to measure each component of its movement within the forest. Two years after the accident, we estimated the annual transportation of (137)Cs from the forest canopy to the floor by litterfall, throughfall and stemflow. Seasonal variations in (137)Cs transportation and differences between forests types were also determined. The total amount of (137)Cs transported from the canopy to the floor in two deciduous and cedar plantation forests ranged between 3.9 and 11.0 kBq m(-2) year(-1). We also observed that (137)Cs transportation with litterfall increased in the defoliation period, simply because of the increased amount of litterfall. (137)Cs transportation with throughfall and stemflow increased in the rainy season, and (137)Cs flux by litterfall was higher in cedar plantation compared with that of mixed deciduous forest, while the opposite result was obtained for stemflow.

OsHKT1;4-mediated Na+ transport in stems contributes to Na+ exclusion from leaf blades of rice at the reproductive growth stage upon salt stress

BackgroundNa+ exclusion from leaf blades is one of the key mechanisms for glycophytes to cope with salinity stress. Certain class I transporters of the high-affinity K+ transporter (HKT) family have been demonstrated to mediate leaf blade-Na+ exclusion upon salinity stress via Na+-selective transport. Multiple HKT1 transporters are known to function in rice (Oryza sativa). However, the ion transport function of OsHKT1;4 and its contribution to the Na+ exclusion mechanism in rice remain to be elucidated.ResultsHere, we report results of the functional characterization of the OsHKT1;4 transporter in rice. OsHKT1;4 mediated robust Na+ transport in Saccharomyces cerevisiae and Xenopus laevis oocytes. Electrophysiological experiments demonstrated that OsHKT1;4 shows strong Na+ selectivity among cations tested, including Li+, Na+, K+, Rb+, Cs+, and NH4+, in oocytes. A chimeric protein, EGFP-OsHKT1;4, was found to be functional in oocytes and targeted to the plasma membrane of rice protoplasts. The level of OsHKT1;4 transcripts was prominent in leaf sheaths throughout the growth stages. Unexpectedly however, we demonstrate here accumulation of OsHKT1;4 transcripts in the stem including internode II and peduncle in the reproductive growth stage. Moreover, phenotypic analysis of OsHKT1;4 RNAi plants in the vegetative growth stage revealed no profound influence on the growth and ion accumulation in comparison with WT plants upon salinity stress. However, imposition of salinity stress on the RNAi plants in the reproductive growth stage caused significant Na+ overaccumulation in aerial organs, in particular, leaf blades and sheaths. In addition, 22Na+ tracer experiments using peduncles of RNAi and WT plants suggested xylem Na+ unloading by OsHKT1;4.ConclusionsTaken together, our results indicate a newly recognized function of OsHKT1;4 in Na+ exclusion in stems together with leaf sheaths, thus excluding Na+ from leaf blades of a japonica rice cultivar in the reproductive growth stage, but the contribution is low when the plants are in the vegetative growth stage.

Development of real-time radioisotope imaging systems for plant nutrient uptake studies

Ionic nutrition is essential for plant development. Many techniques have been developed to image and (or) measure ionic movement in plants. Nevertheless, most of them are destructive and limit the analysis. Here, we present the development of radioisotope imaging techniques that overcome such restrictions and allow for real-time imaging of ionic movement. The first system, called macroimaging, was developed to visualize and measure ion uptake and translocation between organs at a whole-plant scale. Such a device is fully compatible with illumination of the sample. We also modified fluorescent microscopes to set up various solutions for ion uptake analysis at the microscopic level. Both systems allow numerical analysis of images and possess a wide dynamic range of detection because they are based on radioactivity.

Difference in cesium accumulation among rice cultivars grown in the paddy field in Fukushima Prefecture in 2011 and 2012

After the accident of the Fukushima 1 Nuclear Power Plant in March 2011, radioactive cesium was released and paddy fields in a wide area including Fukushima Prefecture were contaminated. To estimate the levels of radioactive Cs accumulation in rice produced in Fukushima, it is crucial to obtain the actual data of Cs accumulation levels in rice plants grown in the actual paddy field in Fukushima City. We herein conducted a two-year survey in 2011 and 2012 of radioactive and non-radioactive Cs accumulation in rice using a number of rice cultivars grown in the paddy field in Fukushima City. Our study demonstrated a substantial variation in Cs accumulation levels among the cultivars of rice.

The effect of fertilization on cesium concentration of rice grown in a paddy field in Fukushima Prefecture in 2011 and 2012

After the accident of the Fukushima 1 nuclear power plant in March 2011, radioactive cesium was released and paddy field in a wide area of Fukushima Prefecture was contaminated. To reduce radioactive Cs uptake by rice, it is important to understand factors that affect Cs uptake in rice. Here we describe our study in 2011 and 2012 to investigate Cs concentration in two rice cultivars, Koshihikari and Hitomebore, the top two cultivars in Fukushima prefecture, grown under different fertilizer conditions in the contaminated paddy field. Our study demonstrated that high nitrogen and low potassium conditions increase Cs concentrations both in straw and brown rice.

Characterization of rapid intervascular transport of cadmium in rice stem by radioisotope imaging

Participation of the intervascular transport system within the rice stem during cadmium (Cd) partitioning was investigated by characterizing 109Cd behaviour in the shoot. In addition, 45Ca, 32P, and 35S partitioning patterns were analysed for comparison with that of 109Cd. Each tracer was applied to the seedling roots for 15min, and the shoots were harvested either at 15min (i.e. immediately after tracer application) or at 48h. Distribution patterns of each element at 15min were studied to identify the primary transport pathway before remobilization was initiated. 32P was preferentially transported to completely expanded leaf blades having the highest transpiration rate. The newest leaf received minimal amounts of 32P. In contrast, the amount of 35S transported to the newest leaf was similar to that transported to the other mature leaf blades. Preferential movement towards the newest leaf was evident for 109Cd and 45Ca. These results directly indicate that elemental transport is differentially regulated in the vegetative stem as early as 15min before the elements are transported to leaves. Cd behaviour in the stem was investigated in detail by obtaining serial section images from the bottom part of shoots after 109Cd was applied to a single crown root. At 30min, the maximum amount of 109Cd was distributed in the peripheral cylinder of the longitudinal vascular bundles (PV) and, interestingly, some amount of 109Cd was transported downwards along the PV. This transport manner of 109Cd provides evidence that Cd can be loaded on the phloem at the stem immediately after Cd is transported from the root.

OsHKT1;5 mediates Na+ exclusion in the vasculature to protect leaf blades and reproductive tissues from salt toxicity in rice

Salt tolerance quantitative trait loci analysis of rice has revealed that the SKC1 locus, which is involved in a higher K+ /Na+ ratio in shoots, corresponds to the OsHKT1;5 gene encoding a Na+ -selective transporter. However, physiological roles of OsHKT1;5 in rice exposed to salt stress remain elusive, and no OsHKT1;5 gene disruption mutants have been characterized to date. In this study, we dissected two independent T-DNA insertional OsHKT1;5 mutants. Measurements of ion contents in tissues and 22 Na+ tracer imaging experiments showed that loss-of-function of OsHKT1;5 in salt-stressed rice roots triggers massive Na+ accumulation in shoots. Salt stress-induced increases in the OsHKT1;5 transcript were observed in roots and basal stems, including basal nodes. Immuno-staining using an anti-OsHKT1;5 peptide antibody indicated that OsHKT1;5 is localized in cells adjacent to the xylem in roots. Additionally, direct introduction of 22 Na+ tracer to leaf sheaths also demonstrated the involvement of OsHKT1;5 in xylem Na+ unloading in leaf sheaths. Furthermore, OsHKT1;5 was indicated to be present in the plasma membrane and found to localize also in the phloem of diffuse vascular bundles in basal nodes. Together with the characteristic 22 Na+ allocation in the blade of the developing immature leaf in the mutants, these results suggest a novel function of OsHKT1;5 in mediating Na+ exclusion in the phloem to prevent Na+ transfer to young leaf blades. Our findings further demonstrate that the function of OsHKT1;5 is crucial over growth stages of rice, including the protection of the next generation seeds as well as of vital leaf blades under salt stress.

Effects of potassium in reducing the radiocesium translocation to grain in rice

Abstract This study investigated the effects of potassium (K) on the behavior of cesium 137 (137Cs) in hydroponically cultured rice plants (Oryza sativa L. “Nipponbare”), revealing that K supply has an influence on reducing 137Cs translocation from leaves to ears as well as decreasing 137Cs uptake by roots. When K was omitted from the culture solution and 137Cs was supplied, the 137Cs content in the ears was increased 6.7-fold compared with the K-sufficient plants, while the total 137Cs content in the shoot part of the K-deficient rice was only 3-fold higher than that in the K-sufficient rice. The fraction of the ears in the total shoot 137Cs content was indeed 2.5 times higher in the K-deficient rice (45%) than that in the K-sufficient rice (18%), demonstrating that K supplementation has an effect on 137Cs translocation to the ears. Irrespective of the K condition, > 95% of the 137Cs that accumulated in the ears was found to be absorbed from the culture solution before ear emergence. K supplementation after ear emergence had an influence on achieving the percentage of 137Cs remobilization in the K-deficient rice close to that in the K-sufficient rice, although the effect on reducing the 137Cs content in the ears could be limited. The results indicate the importance of maintaining an appropriate soil K concentration during the early growth stages to avoid 137Cs contamination of rice grains.

The analysis of magnesium transport system from external solution to xylem in rice root

We report the characterization of magnesium (Mg) transport from external solution to xylem by using xylem fluid analysis in rice (Oryza sativa L. Nipponbare). It was indicated that Mg uptake and/or xylem loading was mediated by two transport systems, a saturable and a linear component. The K m value of the saturable component was approximately 20 µM, and the saturated concentration was calculated to be 1.5 mM. Using a multi-compartment transport box with excised roots, the transfer from external solution to xylem was shown to be inhibited by hexaamminecobalt(III) {[Co(III)(NH3)6], Co-Hex}, a selective inhibitor of CorA-mediated influx of Mg2+, suggesting that the MRS2/MGT genes that belong to the superfamily of CorA-type membrane proteins are involved in the Mg transport system.