Influence of simulated microgravity on physiological reactions in healthy and virus infected wheat plants of different varieties

The effects of clinorotation on the 3 varieties of the wheat plants were studied. The chlorophyll content, photochemical activity of the chloroplasts and changes in wheat streak mosaik virus (WSMV) were analysed. It is established that photosynthetic apparatus in wheat plants, particularly in bred Apogee variety, possesses considerable adaptation potential for the cultivation of plants under the spaceflight conditions.     

Clinostating effects on Apogee wheat resistance to wheat streak mosaic virus

The effects of vertical and horizontal clinostating with container velocity 2 rpm and platform velocity 1 rpm on Apogee wheat resistance to wheat streak mosaic virus (WSMV) were studied. For the first time the yield of grain was obtained from both, healthy and the WSMV-infected wheat plants under conditions of simulated microgravity (clinorotation) which is accounted for by the inclusion of various mechanisms in the process of plant adaptation to the factors of stresses. For the first time the stages of viral infection development were elucidated under the conditions of prolonged clinostating and the presence of the WSMV was detected in wheat plants on the 18th day after inoculation employing the method of polymerase chain reaction (PCR). In the test variant with vertical clinostating (R=1,0) the least favorable conditions for viral reproduction were identified. In the same variant the increases of the total nitrogen content in the ears were observed which may be an evidence of interferon-like protein synthesis in plant similar to the interferon system functioning in animals.     

A model wheat cultivar for transformation to improve resistance to Fusarium Head Blight

Fusarium head blight (FHB), caused primarily by Fusarium graminearum, is a major disease problem in wheat (Triticum aestivum). Genetic engineering holds significant potential to enhance FHB resistance in wheat. Due to the requirement of screening for FHB resistance on flowers at anthesis, the number of screens carried out in a year is limited. Our objective was to evaluate the feasibility of using the rapid-maturing dwarf wheat cultivar Apogee as an alternative genotype for transgenic FHB resistance research. Our transformation efficiency (number of transgenic plants/number of embryos) for Apogee was 1.33%. Apogee was also found to exhibit high FHB susceptibility and reached anthesis within 4 weeks. Interestingly, microsatellite marker haplotype analysis of the chromosome 3BS FHB resistant quantitative trait locus (QTL) region indicated that this region maybe deleted in Apogee. Our results indicate that Apogee is particularly well suited for accelerating transgenic FHB resistance research and transgenic wheat research in general. C.A. Mackintosh and D.F. Garvin contributed equally to the article and should be considered co-first authors     

Clinorotation affects morphology and ethylene production in soybean seedlings

The microgravity environment of spaceflight influences growth, morphology and metabolism in etiolated germinating soybean. To determine if clinorotation will similarly impact these processes, we conducted ground-based studies in conjunction with two space experiment opportunities. Soybean (Glycine max [L.] Merr.) seeds were planted within BRIC (Biological Research In Canister) canisters and grown for seven days at 20 degrees C under clinorotation (1 rpm) conditions or in a stationary upright mode. Gas samples were taken daily and plants were harvested after seven days for measurement of growth and morphology. Compared to the stationary upright controls, plants exposed to clinorotation exhibited increased root length (125% greater) and fresh weight (42% greater), whereas shoot length and fresh weight decreased by 33% and 16% respectively. Plants grown under clinorotation produced twice as much ethylene as the stationary controls. Seedlings treated with triiodo benzoic acid (TIBA), an auxin transport inhibitor, under clinorotation produced 50% less ethylene than the untreated control subjected to the same gravity treatment, whereas a treatment with 2,4-D increased ethylene by five-fold in the clinorotated plants. These data suggest that slow clinorotation influences biomass partitioning and ethylene production in etiolated soybean plants.     

The influence of altered gravity on carbohydrate metabolism in excised wheat leaves

We developed a system to study the influence of altered gravity on carbohydrate metabolism in excised wheat leaves by means of clinorotation. The use of excised leaves in our clinostat studies offered a number of advantages over the use of whole plants, most important of which were minimization of exogenous mechanical stress and a greater amount of carbohydrate accumulation during the time of treatment. We found that horizontal clinorotation of excised wheat leaves resulted in significant reductions in the accumulation of fructose, sucrose, starch and fructan relative to control, vertically clinorotated leaves. Photosynthesis, dark respiration and the extractable activities of ADP glucose pyrophosphorylase (EC 2.7.7.27), sucrose phosphate synthase (EC 2.4.4.14), sucrose sucrose fructosyltransferase (EC 2.4.1.99), and fructan hydrolase (EC 3.2.1.80) were unchanged due to altered gravity treatment.     

Effect of clinorotation on the intensity of lipid peroxidation and superoxide dismutase activity in pea chloroplasts

The lipid peroxidation intensity and the antioxidant enzyme superoxide dismutase (SOD) activity were studied in chloroplasts of Pisum sativum under clinorotation (for 7 and 14 days) for elucidating mechanisms of biological effect of altered gravity. In comparison with the control, increased LP levels in chloroplasts were established during both the terms of clinorotation. The SOD activity rose in the chloroplasts of plants clinorotated for 7 days, which has a significant protective effect. However, under a longer clinorotation (for 14 days) the SOD activity fell, being, however, higher than in the control samples. In accordance with the Selye oxidative stress theory, plants clinorotated for 7 days are in the phase of growing stability, while plants clinorotated for 14 days are in the phase of decreasing stability.     

Fatty acid content and the functional state of pea chloroplasts under altered gravity

The results of the study of the fatty acid content and the functional state of chloroplasts isolated from leaves of pea plants grown during 7 and 14 days in the stationary conditions and under clinorotation (2 rpm) are presented. An increase in the unsaturated fatty acid content occurred after 7-day clinorotation while it insignificantly decreased after more prolonged 14-day clinorotation. A study of the functional state of chloroplasts (the rate of electron transport in photosystems II [PSII] and in photosystem I [PSI] and in the whole photosynthetic electron transport chain) showed its decrease under both terms of clinorotation in comparison with control ones. In addition, 14-day clinorotation caused more significant lowering of the electron transport rate, particularly in PSI. Changes in both the fatty acid content and the electron transport rate are discussed in relation to the activation of lipid peroxidation and the increased production of activated oxygen species in chloroplasts under clinorotation.     

Clinorotation effect on thermodynamic efficiency of energy transformation in chloroplasts of higher plants

The aim of the work was to estimate the effect of clinorotation on thermodynamic coupling and efficiency of the process of photosynthetic energy transformation in chloroplasts using nonequilibrium thermodynamics approach. These parameters were calculated from experimentally determined net photosynthetic oxygen evolution at static head [(Je)sh], uncoupled rate of the oxygen evolution (Jo)unc and net rate of ATP production. It was found that in chloroplasts isolated from control and clinorotated pea plants coefficient of thermodynamic coupling of photophosphorylation (q) was 0.94 and 0.91 respectively. Optimal thermodynamic efficiency (eta opt) of the systems were calculated as 0.64 and 0.6 for control and clinorotation plants. Thus the data of the work show that thermodynamic efficiency of light energy transformation in higher plants is under influence of imitated weightlessness conditions.     

Ultrastructure of statocytes and cells of distal elongation zone of Arabidopsis thaliana under clinorotation

Results of the electron-microscopic investigation of root apices of Arabidopsis thaliana 3-, 5- and 7-days-old seedlings grown in the stationary conditions and under clinorotation are presented. It was shown the similarity in the root apex cell ultrastructure in control and under clinorotation. At the same time there were some differences in the ultrastructure of statocytes and the distal elongation zone under clinorotation. For the first time the sensitivity of ER-bodies, which are derivatives of GER and contain beta-glucosidase, to the influence of simulated microgravity was demonstrated by increased quantity and area of ER-bodies at the cell section as well as by higher variability of their form under clinorotation. A degree of these changes correlated with the duration of clinorotation. On the basis of experimental data a protective role of ER-bodies in adaptation of plants to microgravity is supposed.     

Protein expression in Arabidopsis thaliana after chronic clinorotation

Soluble protein expression in Arabidopsis thaliana L. (Heynh.) leaf and stem tissue was examined after chronic clinorotation. Seeds of Arabidopsis were germinated and plants grown to maturity on horizontal or vertical slow-rotating clinostats (1 rpm) or in stationary vertical control units. Total soluble proteins and in vivo-labeled soluble proteins isolated from these plants were analyzed by two-dimensional SDS PAGE and subsequent fluorography. Visual and computer analysis of the resulting protein patterns showed no significant differences in either total protein expression or in active protein synthesis between horizontal clinorotation and vertical controls in the Arabidopsis leaf and stem tissue. These results show chronic clinorotation does not cause gross changes in protein expression in Arabidopsis.     

Protein patterns of the Brassica rapa ovules and seeds under altered gravity

Electrophoretic investigation of protein patterns of Brassica rapa L. ovules and seeds from plants grown under clinorotation and in the laboratory control was carried out. Ovules at different stages (7 and 18 days after pollination) and mature seeds were analyzed. Polymorphism of seed storage proteins of B. rapa was taken into consideration in analysis of changes in ovule protein patterns under clinorotation. The appearance of a protein component in the region of about 43 kDa was detected in protein patterns of 7-day-old and 18-day-old ovules in the clinostat variants. Under altered gravity, in 18-day-old ovules, the appearance of a protein in the region of about 70 kDa was also revealed. The appearance of the protein component with the similar mobility (about 43 kDa) in ovules of different age from plants grown at clinorotation suggests that synthesis of this protein may be associated with the plant response to altered gravity. However, the investigation of the nature of this protein and its role requires further research to rule out its appearance because of genotypic differences between ovules of the control and experimental variants.     

Growth,yield and reproduction of dwarf tomato grown under simulated microgravity conditions

Abstract

A closed hydroponic system combined with a horizontal uniaxial clinostat has been used to grow tomato plants (Solanum lycopersicum L.) under simulated microgravity conditions. The study was carried out to evaluate the quanti-qualitative traits (growth, yield and quality) of the dwarf tomato variety ‘Micro-Tom’ grown under simulated microgravity conditions and to determine if tomato plants would complete their life cycle (‘seed-to-seed’). Morphological and growth characteristics of ‘Micro-Tom’ were modified during clinorotation treatment. The ‘Micro-Tom’ plants grown under simulated microgravity exhibited a spreading growth and an increasing of the internode length. Total fruit yield, small fruit yield, leaf area, leaf dry weight, fruit dry weight, total dry weight and shoot – root ratio were lower in the clinorotated tomato plants than those grown in the control treatment. Foliar amount of carotenoids, and chlorophyll a and b were also substantially reduced under simulated microgravity conditions. Quality parameters (total soluble solids and fruit dry matter) of tomato plants were also negatively affected by clinorotation. The number of flowers per plant was increased by 32% in clinorotated plants versus controls. Fruit setting was reduced by 46% under clinorotation, while no significant difference was recorded for the pollen fertility and the seed number in small and large fruits. Clinorotation-exposed and control seeds were used in a germination trial in order to evaluate whether the seeds so formed were viable and if subsequent generations might be obtained in microgravity. Seeds formed under simulated microgravity proved to be biologically and functionally complete (germination = 78.6%) showing that ‘Micro-Tom’ plants could realize complete ontogenesis, from seed to seed in microgravity.     

Root-hair response to vector-free gravity: role of Ca2+ ions

In plants, apical growth is demonstrated by a variety of cells, including root hairs (RH) which are tubular outgrowths of root epidermal cells. They are likely to be involved in uptake of nutrients and water, anchorage of plants, maintenance of contact between roots and soil, and root exudation. Over the last years, it has become clear that calcium is involved in various processes which result in tip growth. Previous studies from our laboratory have demonstrated an increase in calcium level in root cells of pea seedlings grown under conditions of space flight and clinorotation. On the basis of these data, we have suggested that such effects of microgravity and clinorotation might be due to the enhancement of Ca2+ influx into hyaloplasm evidently through Ca2+ channels. In this regard, it was interesting to examine the effects of clinorotation and Ca2+ channel blockers (verapamil and nifedipine) on orientation and structure of growing RH that might be an appropriate model system being sensitive to calcium for studying the gravitational effects at cellular level.     

Role of cytoskeleton in gravisensing of the root elongation zone in Arabidopsis thaliana plants

In order to reveal the involvement of tubulin microtubules and actin microfilaments in gravisensing reactions in the distal elongation zone of root, Arabidopsis thaliana plants stably transformed with MAP4-GFP construct were grown under slow clinorotation. Experiments have shown that stabilization of cell growth in the distal elongation zone of Arabidopsis seedling root is provided by common structural organization of microtubules and microfilaments, and interrelations between microtubules and microfilaments is highly dependent upon the type of cell differential growth. Less pronounced effect of microfilament disruption on microtubule organization has been observed under clinorotation and it suggests the existence of complex mechanism of cooperation between microtubules and microfilaments which is probably, masked on earth.     

禾谷多粘菌对小麦梭条斑花叶病毒的传播及其土壤侵染潜力的测定

从大田侵染小麦梭条斑花叶病毒的小麦病根中挑取禾谷多粘菌休眠孢子堆,接种受侵染小麦品种扬麦４号,经砂培养纯化,获得５个禾谷多粘菌分离物,但都为无毒。无毒多粘菌休眠孢子堆接种表现ＷＳＳＭＶ症状的小麦,经培养可饲获病毒,并可经接咱后将病毒传播给无病小麦,供试的４个大小麦禾谷多粘菌分离物都可对大小进行交叉侵染,产生同样数量的游动孢子产量。供试５个病土和２个无病土样品,都具有强大持多粘菌侵染潜力,即使稀释放     

The effect of clinorotation on vestibular compensation in upside-down swimming catfish.

Upside-down swimming catfish Synodontis nigriventris can keep upside-down swimming posture stably under pseudo-microgravity generated by clinostat. When the vestibular organ is unilaterally ablated, the operated S. nigriventris shows disturbed swimming postures under the clinorotation condition. However, about 1 month after the operation, unilateral vestibular organ-ablated S. nigriventris shows stable upside-down swimming posture under the condition (vestibular compensation). In contrast, a closely related upside-up swimming catfish Synodontis multipunctatus belonging to same Synodontis family can not keep stable swimming postures under the clinorotation conditions. In this study, we examined the effect of continuous clinorotation on vestibular compensation in intact and unilateral vestibular organ-ablated Synodontis nigriventris and Synodontis multipunctatus. After the exposure to continuous clinorotation, the postures of the catfish were observed under microgravity provided by parabolic flights of an aircraft. Unilateral vestibular organ-ablated S. nigriventris which had been exposed to continuous clinorotation showed stable swimming postures and did not show dorsal light reaction (DLR) under microgravity. This postural control pattern of the operated catfish was similar to that of intact catfish. Intact and unilateral vestibular organ-ablated S. multipunctatus showed DLR during microgravity. Our results confirmed that S. nigriventris has a novel balance sensation which is not affected by microgravity. DLR seems not to play an important role in postural control. It remains unclear that the continuous clinorotation effects on vestibular compensation because we could not keep used unilateral vestibular organ-ablated fish alive under continuous clinorotation for uninterrupted 25 days. This study suggests that space flight experiments are required to explore whether gravity information is essential for vestibular compensation.     

Structural and cytochemical aspects of <Emphasis Type="Italic">Brassica rapa</Emphasis> L. embryogenesis under clinorotation

The results of study of embryo development in B. rapa plants as well as rate and character of nutrient substance accumulation in their cells under slow horizontal clinorotation and the laboratory control were presented. Significant similarity in the peculiarities of embryo differentiation and character of nutrient substance accumulation in both variants was established. The cases of different deviations during embryo differentiation, quantity of reserve nutrient substances, and the rate of their accumulation in the cells were revealed under clinorotation in comparison with the laboratory control.     

Effects of mechanostimulation on gravitropism and signal persistence in flax roots

Gravitropism describes curvature of plants in response to gravity or differential acceleration and clinorotation is commonly used to compensate unilateral effect of gravity. We report on experiments that examine the persistence of the gravity signal and separate mechanostimulation from gravistimulation. Flax roots were reoriented (placed horizontally for 5, 10 or 15 min) and clinorotated at a rate of 0.5 to 5 rpm either vertically (parallel to the gravity vector and root axis) or horizontally (perpendicular to the gravity vector and parallel to the root axis). Image sequences showed that horizontal clinorotation did not affect root growth rate (0.81 ± 0.03 mm h⁻¹) but vertical clinorotation reduced root growth by about 7%. The angular velocity (speed of clinorotation) did not affect growth for either direction. However, maximal curvature for vertical clinorotation decreased with increasing rate of rotation and produced straight roots at 5 rpm. In contrast, horizontal clinorotation increased curvature with increasing angular velocity. The point of maximal curvature was used to determine the longevity (memory) of the gravity signal, which lasted about 120 min. The data indicate that mechanostimulation modifies the magnitude of the graviresponse but does not affect memory persistence.Key words: mechanostimulation, memory, clinorotation speed and direction, signal persistence, signal saturation     

Microgravity and clinorotation cause redistribution of free calcium in sweet clover columella cells

In higher plants, calcium redistribution is believed to be crucial for the root to respond to a change in the direction of the gravity vector. To test the effects of clinorotation and microgravity on calcium localization in higher plant roots, sweet clover (Melilotus alba L.) seedlings were germinated and grown for two days on a slow rotating clinostat or in microgravity on the US Space Shuttle flight STS-60. Subsequently, the tissue was treated with a fixative containing antimonate (a calcium precipitating agent) during clinorotation or in microgravity and processed for electron microscopy. In root columella cells of clinorotated plants, antimonate precipitates were localized adjacent to the cell wall in a unilateral manner. Columella cells exposed to microgravity were characterized by precipitates mostly located adjacent to the proximal and lateral cell wall. In all treatments some punctate precipitates were associated with vacuoles, amyloplasts, mitochondria, and euchromatin of the nucleus. A quantitative study revealed a decreased number of precipitates associated with the nucleus and the amyloplasts in columella cells exposed to microgravity as compared to ground controls. These data suggest that roots perceive a change in the gravitational field, as produced by clinorotation or space flights, and respond respectively differently by a redistribution of free calcium.     

Life history of the bird cherry-oat aphid, Rhopalosiphum padi (Homoptera: Aphididae), on transgenic and untransformed wheat challenged with barley yellow dwarf virus

The life history of Rhopalosiphum padi (L.) was monitored on transgenic and untransformed (soft white winter wheat plants that were infected with Barley yellow dwarf virus (BLDV), noninfected, or challenged with virus-free aphids under laboratory conditions. Two transgenic soft white winter wheat genotypes (103.1J and 126.02) derived from the parental variety Lambert and expressing the barley yellow dwarf virus coat protein gene, and two untransformed varieties, virus-susceptible Lambert and virus-tolerant Caldwell, were tested. B. padi nymphal development was significantly longer on the transgenic genotypes infected with BYDV, compared with noninfected transgenic plants. In contrast, nymphal development on Lambert was significantly shorter on BYDV-infected than on noninfected plants. Nymphal development on noninfected Lambert was significantly longer than on noninfected transgenics. No significant difference in nymphal development period was detected between virus-infected and noninfected Caldwell. Aphid total fecundity, length of reproductive period, and intrinsic rate of increase were significantly reduced on BYDV-infected transgenic plants compared with BYDV-infected Lambert. In contrast, reproductive period, total adult fecundity, and intrinsic rate of increase on noninfected Lambert were significantly reduced compared with noninfected transgenics. Transgenic plants infected with BYDV were inferior hosts for R. padi compared with infected Lambert. However, noninfected transgenics were superior hosts for aphids than noninfected Lambert. Moderate resistance to BYDV, as indicated by a significantly lower virus titer, was detected in the transgenic genotypes compared with the untransformed ones. Results show for the first time that transgenic virus resistance in wheat can indirectly influence R. padi life history.