Positron-emitting Tracer Imaging System Research Articles

Rice metal-nicotianamine transporter, OsYSL2, is required for the long-distance transport of iron and manganese

Rice (Oryza sativa) is indispensable in the diet of most of the world's population. Thus, it is an important target in which to alter iron (Fe) uptake and homeostasis, so as to increase Fe accumulation in the grain. We previously isolated OsYSL2, a functional iron [Fe(II)]- and manganese [Mn(II)]-nicotianamine complex transporter that is expressed in phloem cells and developing seeds. We produced RNAi (OsYSL2i) and overexpression lines (OXOsYSL2) of OsYSL2. At the vegetative stage in an OsYSL2i line, the Fe and Mn concentrations were decreased in the shoots, and the Fe concentration was increased in the roots. At the reproductive stage, positron-emitting tracer imaging system analysis revealed that Fe translocation to the shoots and seeds was suppressed in OsYSL2i. The Fe and Mn concentrations were decreased in the seeds of OsYSL2i, especially in the endosperm. Moreover, the Fe concentration in OXOsYSL2 was lower in the seeds and shoots, but higher in the roots, compared with the wild type. Furthermore, when OsYSL2 expression was driven by the sucrose transporter promoter, the Fe concentration in the polished rice was up to 4.4-fold higher compared with the wild type. These results indicate that the altered expression of OsYSL2 changes the localization of Fe, and that OsYSL2 is a critical Fe-nicotianamine transporter important for Fe translocation, especially in the shoots and endosperm.

Real-time Analysis of Translocation of Photosynthates to Nodules in Soybean Plants Using 11CO2 with a Positron-emitting Tracer Imaging System(PETIS)

Real-time Analysis of Translocation of Photosynthates to Nodules in Soybean Plants Using 11CO2 with a Positron-emitting Tracer Imaging System(PETIS)

Determination of the rate of 18F-labeled water movement to the leaf and its association with water relations in acid soil-tolerant rice varieties

The visual symptoms of damage by acid sulfate soil look quite similar to those due to soil moisture deficit. The present paper is to confirm if acid soil-tolerance is associated with drought tolerance in terms of leaf water potential, transpiration, and dry matter production. Seedlings of seven acid soil-tolerant (acid-tolerant), and three acid soil-intolerant varieties (acid-intolerant) were subjected to low pH conditions (pH 3.8) for 48 hours. The rate of water uptake was determined continuously by measuring radioactivity in the collar part (target area) of rice leaves exposed to 18F-labeled water (18F-water) using a Positron Emitting Tracer Imaging System (PETIS). The PETIS measurement showed that the radioactivity in the target position of leaves of acid-tolerant varieties increased faster than that of the intolerant varieties after the 18F-water was applied into the vial. In the plants subjected to low pH conditions, the transpiration rate (Tr) and leaf water potential (Ψ) decreased in both acid-tolerant and intolerant varieties. However, the extent of reduction was less in the acid-tolerant varieties than in the intolerant ones. Moreover, the dry matter production rate of the acid-tolerant varieties was significantly higher than that of the acid-intolerant ones in the low pH conditions. This study clearly shows that 18F-water uptake in a leaf was higher, and water relations were also better maintained in the acid-tolerant varieties compared with intolerant ones, resulting in higher growth rate in the acid-tolerant varieties, when plants were exposed to the acid solution conditions. We conclude that acid soil-tolerance is closely associated with leaf water relations in rice plants.

Real-time imaging of nitrogen fixation in an intact soybean plant with nodules using 13N-labeled nitrogen gas

Real-time images of nitrogen fixation in an intact nodule of hydroponically cultured soybean (Glycine max [L] Merr.) were obtained. In the present study, we developed a rapid method to produce and purify 13N-labeled radioactive nitrogen gas (half life: 9.97 min). 13N was produced from a 16O (p, α) 13N nuclear reaction. The target chamber was filled with CO2 and irradiated for 10 min with protons at an energy of 18.3 MeV and an electric current of 5 μA, which was delivered from a cyclotron. All CO2 in the collected gas was absorbed and removed with powdered soda-lime in a syringe and replaced with helium gas. The resulting gas was injected into gas chromatography and separated and a 35 mL fraction, including the peak of [13N]-nitrogen gas, was collected by monitoring the chromatogram. The obtained gas was mixed with 10 mL of O2 and 5 mL of N2 and used in the tracer experiment. The tracer gas was fed into the underground part of intact nodulated soybean plants and serial images of the distribution of 13N were obtained non-invasively using a positron-emitting tracer imaging system (PETIS). The rates of nitrogen fixation of the six test plants were estimated to be 0.17 ± 0.10 μmol N2 h−1 from the PETIS image data. The decreasing rates of assimilated nitrogen were also estimated to be 0.012 ± 0.011 μmol N2 h−1. In conclusion, we successfully observed nitrogen fixation in soybean plants with nodules non-invasively and quantitatively using [13N]N2 and PETIS.

Production of no-carrier-added 64Cu and applications to molecular imaging by PET and PETIS as a biomedical tracer

Copper-64 was produced by the 64Ni(p, n)64Cu reaction using enriched 64NiO target. We investigated and compared the production yield of 64Cu for proton beams of various energies by using a thick target. Enriched 64Ni was recovered with high yield by simple procedures. Imaging studies using positron emission tomography (PET) and positron emitting tracer imaging system (PETIS) were performed. We obtained clear images in PET and PETIS studies. The results of this study indicate that 64Cu can be utilized as a biomedical tracer for the molecular imaging both in animals and plants.

52Fe Translocation in Barley as Monitored by a Positron-Emitting Tracer Imaging System (PETIS): Evidence for the Direct Translocation of Fe from Roots to Young Leaves via Phloem

The real-time translocation of iron (Fe) in barley (Hordeum vulgare L. cv. Ehimehadaka no. 1) was visualized using the positron-emitting tracer 52Fe and a positron-emitting tracer imaging system (PETIS). PETIS allowed us to monitor Fe translocation in barley non-destructively under various conditions. In all cases, 52Fe first accumulated at the basal part of the shoot, suggesting that this region may play an important role in Fe distribution in graminaceous plants. Fe-deficient barley showed greater translocation of 52Fe from roots to shoots than did Fe-sufficient barley, demonstrating that Fe deficiency causes enhanced 52Fe uptake and translocation to shoots. In the dark, translocation of 52Fe to the youngest leaf was equivalent to or higher than that under the light condition, while the translocation of 52Fe to the older leaves was decreased, in both Fe-deficient and Fe-sufficient barley. This suggests the possibility that the mechanism and/or pathway of Fe translocation to the youngest leaf may be different from that to the older leaves. When phloem transport in the leaf was blocked by steam treatment, 52Fe translocation from the roots to older leaves was not affected, while 52Fe translocation to the youngest leaf was reduced, indicating that Fe is translocated to the youngest leaf via phloem in addition to xylem. We propose a novel model in which root-absorbed Fe is translocated from the basal part of the shoots and/or roots to the youngest leaf via phloem in graminaceous plants.

Use of positron-emitting tracer imaging system for measuring the effect of salinity on temporal and spatial distribution of 11C tracer and coupling between source and sink organs

Salinity stress affects photosynthate partitioning between sources and sinks of plants, but how it affects these systems is less well understood. Because sources and sinks are closely tied, any adverse effect under suboptimal conditions on one of these is often misinterpreted for an effect on the other. Carbon partitioning is indispensable for stress resistance and good plant growth. In the present study, carbon partitioning in tomato plants ( Lycopersicon esculentum L. cv. Momotarou) in a saline (NaCl) environment was studied by feeding radioactive 11C and stable 13C isotopes. Pulse-chases were conducted to measure the spatial and temporal distribution of 13C. 13C was measured by a standard conventional technique, but 11C distribution was monitored using a positron-emitting tracer imaging system (PETIS). Salt stress resulted in reduced carbon translocation toward roots. The majority of the photosynthate accumulated in the leaf. We also observed that the reduction in translocation of carbon occurred well before the salt stress symptoms of reduced photosynthesis and reduced plant growth in salt-exposed plants. The effect on sink activity was also shown by a decrease in stem diameter. In addition, PETIS analysis of 11C translocation indicated that carbon translocation to roots was inhibited under salt conditions without a direct effect on leaf Na accumulation or osmotic stress. These results suggest that NaCl has direct effects on plants, inhibiting carbon partitioning within a few hours of salt exposure without inhibition of source activity.

Analysis of NO3 interception of the parasitic angiosperm Orobanche spp. using a positron-emitting tracer imaging system and 13NO− 3: A new method for the visualization and quantitative analysis of the NO3 interception ratio

The root parasitic plants Orobanche spp. (broomrapes) seriously affect agricultural production. A visualization and quantitative analytical method for the interception of nutrients was established using a positron-emitting tracer imaging system and 13NO− 3. By using this analytical method that involves volume normalization with 18F− images, the nitrogen nutrient interception ratio of the Orobanche spp. was calculated to be 73.6 ± 3.9% in a host–parasite system of red clover (Trifolium pratense L.).

Deoxymugineic acid increases Zn translocation in Zn-deficient rice plants

Deoxymugineic acid (DMA) is a member of the mugineic acid family phytosiderophores (MAs), which are natural metal chelators produced by graminaceous plants. Rice secretes DMA in response to Fe deficiency to take up Fe in the form of Fe(III)–MAs complex. In contrast with barley, the roots of which secrete MAs in response to Zn deficiency, the amount of DMA secreted by rice roots was slightly decreased under conditions of low Zn supply. There was a concomitant increase in endogenous DMA in rice shoots, suggesting that DMA plays a role in the translocation of Zn within Zn-deficient rice plants. The expression of OsNAS1 and OsNAS2 was not increased in Zn-deficient roots but that of OsNAS3 was increased in Zn-deficient roots and shoots. The expression of OsNAAT1 was also increased in Zn-deficient roots and dramatically increased in shoots; correspondingly, HPLC analysis was unable to detect nicotianamine in Zn-deficient shoots. The expression of OsDMAS1 was increased in Zn-deficient shoots. Analyses using the positron-emitting tracer imaging system (PETIS) showed that Zn-deficient rice roots absorbed less 62Zn-DMA than 62Zn2+. Importantly, supply of 62Zn-DMA rather than 62Zn2+ increased the translocation of 62Zn into the leaves of Zn-deficient plants. This was especially evident in the discrimination center (DC). These results suggest that DMA in Zn-deficient rice plants has an important role in the distribution of Zn within the plant rather than in the absorption of Zn from the soil.Electronic supplementary materialThe online version of this article (doi:10.1007/s11103-008-9292-x) contains supplementary material, which is available to authorized users.

Real-time Analysis of Photoassimilate Translocation in Intact Eggplant Fruit using 11CO2 and a Positron-emitting Tracer Imaging System

A recently developed positron-emitting tracer imaging system (PETIS) noninvasively produces quantitative real-time images of the movement of various compounds in plants. To clarify the mechanism of the growth and development of the fruit of eggplant (Solanum melongena L.), we fed 11CO2 to a leaf and monitored the translocation of 11C-labeled photoassimilate into the fruit by PETIS. Continuous images of the translocation of [11C]photoassimilate from the leaf to the fruit and inside the fruit were produced, with a shorter time resolution than reported previously. 11C signal intensity in the fruit increased gradually, and its distribution was not uniform. The fruits were sliced after PETIS measurement and exposed to the imaging plates of a bio-imaging analyzer system. The resultant images showed the localization of 11C activity inside the fruit, and indicated that translocation of photoassimilate within the fruit was not uniform. The velocity of photoassimilate translocation in the peduncle and changes in the rate of translocation of photoassimilate in the fruit were estimated by analysis of PETIS data. The velocity of photoassimilate translocation through the peduncle was estimated as 1.17 cm·min−1. About 60 min after the start of 11CO2 feeding, the 11C activity of the fruit began to increase, and by 120 min it had reached about 8% of feeding 11CO2. These results indicate that it took about 60 min for the first [11C]photoassimilates to reach the fruit. Real time observation of photoassimilate translocation within a fruit has never been reported. PETIS may be a powerful tool for revealing the mechanisms of fruit development and maturity.

A Preliminary Study of an Al Effect on 15O-Labeled Water Uptake in a Soybean Plant by PETIS (Positron Emitting Tracer Imaging System)

A Preliminary Study of an Al Effect on 15O-Labeled Water Uptake in a Soybean Plant by PETIS (Positron Emitting Tracer Imaging System)

52Mn translocation in barley monitored using a positron-emitting tracer imaging system

Until now, the real-time uptake and movement of manganese (Mn), an essential plant nutrient, has not been documented in plants. In this study, the real-time translocation of Mn in barley (Hordeum vulgare L. cv. Ehimehadaka no. 1) was visualized using the positron-emitting tracer 52Mn and a positron-emitting tracer imaging system (PETIS). PETIS allowed the non-destructive monitoring of Mn translocation in barley under various conditions. In all cases, 52Mn first accumulated in the discrimination center (DC) at the basal portion of the shoot, suggesting that this region may play an important role in Mn distribution in graminaceous plants. Manganese-deficient barley showed greater translocation of 52Mn from roots to shoots than did Mn-sufficient barley, demonstrating that Mn deficiency causes enhanced Mn uptake and loading into vascular bundles. In contrast, the translocation of 52Mn from roots to shoots was suppressed in Mn-excess barley. In these plants, the uptake of Mn may be suppressed or Mn may accumulate in the intercellular organelles of root cells, resulting in low rates of Mn translocation to shoots. In Mn-sufficient barley, the dark treatment did not suppress the translocation of 52Mn to the youngest leaf, suggesting that the translocation of Mn to the youngest leaf is independent of the transpiration stream. When 52Mn was supplied to the cut end of an expanded leaf, 52Mn was transported to the DC within 27 min and then retranslocated to roots and other leaves. Our results show that the translocation of Mn from the roots to the DC depends passively on water flow, but actively on the Mn transporter(s).

Kinetic Analysis of Carbon-11-Labeled Carbon Dioxide for Studying Photosynthesis in a Leaf Using Positron Emitting Tracer Imaging System

The positron emitting tracer imaging system (PETIS) and carbon-11-labeled carbon dioxide (11CO2) can be used for imaging the photosynthesis process in plant leaves. Further, 11C kinetics facilitate the estimation of the physiological function parameters of photosynthesis. PETIS measurements were performed under four light conditions for each exposure of a single leaf to 11CO2 gas. In order to estimate the rate constants of the photosynthesis parameters, the time-activity curves of the input 11CO2 gas and the leaf response were fitted to an appropriate compartmental tracer kinetic model that considers photoassimilation and sucrose export rate constants as influx and efflux, respectively. The data obtained by this method show a reasonable response with respect to the photoenvironment of the leaf, and they are important for discussing photosynthesis with regard to plant physiology and agriculture

Biosynthesis and secretion of mugineic acid family phytosiderophores in zinc‐deficient barley

Mugineic acid family phytosiderophores (MAs) are metal chelators that are produced in graminaceous plants in response to iron (Fe) deficiency, but current evidence regarding secretion of MAs during zinc (Zn) deficiency is contradictory. Our studies using HPLC analysis showed that Zn deficiency induces the synthesis and secretion of MAs in barley plants. The levels of the HvNAS1, HvNAAT-A, HvNAAT-B, HvIDS2 and HvIDS3 transcripts, which encode the enzymes involved in the synthesis of MAs, were increased in Zn-deficient roots. Studies of the genes involved in the methionine cycle using microarray analysis showed that the transcripts of these genes were increased in both Zn-deficient and Fe-deficient barley roots, probably allowing the plant to meet its demand for methionine, a precursor in the synthesis of MAs. In addition, HvNAAT-B transcripts were detected in Zn-deficient shoots, but not in those that were deficient in Fe. Increased synthesis of MAs in Zn-deficient barley was not due to a deficiency of Fe, because Zn-deficient barley accumulated more Fe than did the control plants, ferritin transcripts were increased in Zn-deficient plants, and Zn deficiency promoted Fe transport from root to shoot. Moreover, analysis using the positron-emitting tracer imaging system (PETIS) confirmed that more 62Zn(II)-MAs than 62Zn2+ were absorbed by the roots of Zn-deficient barley plants. These data suggest that the increased biosynthesis and secretion of MAs arising from a shortage of Zn are not due to an induced Fe deficiency, and that secreted MAs are effective in absorbing Zn from the soil.

Rice plants take up iron as an Fe3+‐phytosiderophore and as Fe2+

Only graminaceous monocots possess the Strategy II iron (Fe)-uptake system in which Fe is absorbed by roots as an Fe3+-phytosiderophore. In spite of being a Strategy II plant, however, rice (Oryza sativa) contains the previously identified Fe2+ transporter OsIRT1. In this study, we isolated the OsIRT2 gene from rice, which is highly homologous to OsIRT1. Real-time PCR analysis revealed that OsIRT1 and OsIRT2 are expressed predominantly in roots, and these transporters are induced by low-Fe conditions. When expressed in yeast (Saccharomyces cerevisiae) cells, OsIRT2 cDNA reversed the growth defects of a yeast Fe-uptake mutant. This was similar to the effect of OsIRT1 cDNA. OsIRT1- and OsIRT2-green fluorescent protein fusion proteins localized to the plasma membrane when transiently expressed in onion (Allium cepa L.) epidermal cells. OsIRT1 promoter-GUS analysis revealed that OsIRT1 is expressed in the epidermis and exodermis of the elongating zone and in the inner layer of the cortex of the mature zone of Fe-deficient roots. OsIRT1 expression was also detected in the ccompanion cells. Analysis using the positron-emitting tracer imaging system showed that rice plants are able to take up both an Fe3+-phytosiderophore and Fe2+. This result indicates that, in addition to absorbing an Fe3+-phytosiderophore, rice possesses a novel Fe-uptake system that directly absorbs the Fe2+, a strategy that is advantageous for growth in submerged conditions.

A new visualization technique for the study of the accumulation of photoassimilates in wheat grains using [ 11C]CO 2

Non-invasive real-time visualization of the accumulation of photoassimilates in the grains of an ear of wheat using [ 11C]CO 2 and positron emitting tracer imaging system (PETIS) was studied. [ 11C]CO 2 was supplied to the center of a fully expanded leaf of a wheat plant for an initial 10 min, and the transportation of 11C-labeled photoassimilates into the grains of the ear was monitored for 120 min using the PETIS. Each grain was clearly identified in the obtained animation. The 11C-labeled photoassimilates arrived at the ear from the [ 11C]CO 2-absorbing leaf within 53 min from the time of supplying [ 11C]CO 2. After that, grains appeared on the image one by one from the basal part and full images of the grains appeared within 20 min. The time course of the accumulation of photoassimilates into each grain showed a different profile. Furthermore, the PETIS data suggested that the photo-condition of the ear plays an important role in the transportation of photoassimilates in wheat. PETIS can be used to visualize the dynamics of the substances in a living plant in real time and can exhibit the time course analysis of substances, such as the transportation, distribution, and accumulation.

Quantitative Modeling of Photoassimilate Flow in an Intact Plant Using the Positron Emitting Tracer Imaging System (PETIS)

The photoassimilate flow in an intact plant stem was imaged in real-time and its dynamics was quantitatively described using the Positron Emitting Tracer Imaging System (PETIS). Radioactive 11CO2 was fed to a leaf of an intact broad bean (Vicia faba L.) plant, together with air containing an ambient concentration of non-radioactive carrier CO2 gas. Movies of flow of the 11C-labeled photoassimilates in the plant body were captured with PETIS. Here we demonstrate that the average flow speeds and the distribution ratios of photoassimilates in the respective nodes and internodes of the observed stem can be estimated by the transfer function analysis, one of the mathematical modeling methods. We also estimated the changes in the spatial distribution of the average flow speeds in the same stem when the fed leaf was exposed to enriched carrier CO2 gas.

Input–output analysis of in vivo photoassimilate translocation using Positron-Emitting Tracer Imaging System (PETIS) data

The Positron-Emitting Tracer Imaging System (PETIS) is introduced for monitoring the distribution of (11)C-labelled photoassimilates in Sorghum. The obtained two-dimensional image data were quantitatively analysed using a transfer function analysis approach. While one half of a Sorghum root in a split root system was treated with either 0, 100, or 500 mM NaCl dissolved in the nutrient solution, tracer images of the root halves and the lower stem section were recorded using PETIS. From the observed tracer levels, parameters were estimated, from which the mean speed of tracer transport and the proportion of tracer moved between specified image positions were deduced. Transport speed varied between 0.7 and 1.8 cm min(-1) with the difference depending on which part of the stem was involved. When data were collected in the lowest 0.5-1 cm of the stem, which included the point where the roots emerge, transport speed was less. Rapid changes in NaCl concentration, from 0 to 100 mM, resulted in short-term increases of assimilate import into the treated root. This response represented a transient osmotic effect, that was compensated for in the medium-term by osmotic adaptation. Higher concentrations of NaCl (500 mM) resulted in distinctly less photoassimilate transport into the treated root half. The present results agree with earlier observations, showing that transport of (11)C-labelled photoassimilates measured with the PETIS detector system can be quantified using the method of input-output analysis. It is worth noting that with the PETIS detector system, areas of interest do not need to be defined until after data collection. This means that unexpected behaviour of a plant organ will be seen, which is not necessarily the case with conventional detector systems looking at predefined areas of interest.

Positron Emitting Tracer Imaging System (PETIS)によって観測した植物体内におけるトレーサー移動の動画像データの数理的解析

Positron Emitting Tracer Imaging System (PETIS)によって観測した植物体内におけるトレーサー移動の動画像データの数理的解析

H2 15O translocation in rice was enhanced by 10 μm 5-aminolevulinic acid as monitored by positron emitting tracer imaging system (PETIS)

5-Aminolevulinic acid (ALA) acts to increase chlorophyll biosynthesis, photosynthesis, cold stress tolerance, and salt tolerance at low concentrations. We studied the effects of ALA on H2 15O translocation from the roots to the shoots of rice plants (Oryza sativa L. cv. Nipponbare) in real time by a positron-emitting tracer imaging system (PETIS). When the plant was treated with 10 μm ALA, the velocity of the H2 15O translocation from 2 to 12 min after absorption increased to 126, 137, 140% that of the control at 1.5, 2.5, and 3.5 h after ALA treatment, respectively. However, ALA did not affect the H2 15O translocation within 0.5 h of treatment. When the plant was treated with 0.1 mM ABA at 4 h after 10 μm ALA treatment, the velocity of the H2 15O translocation decreased at 0.5 h after ABA treatment. Those observations suggested ALA might be absorbed and transported to the guard cells within 1.5 h and functioned to expand the stomatal aperture.