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.     

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 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 effect of simulated microgravity on formation of the pigment apparatus in etiolated barley seedlings

The effect of horizontal clinorotation on the dynamics of the accumulation of the main photosynthetic pigments in the greening of 6-day-old etiolated barley seedlings has been studied. The content of protochlorophillide, the direct precursor of chlorophyll a, in clinorotated seedlings in the dark was 9–20% lower than in the control group. After exposure of barley seedlings to light for 12 h under clinorotation, chlorophyll accumulation lagged behind the control by 45% and reached the control value after 48–72 h. The total content of carotenoids increased many fold during greening; at the first stage the carotenoid level in clinorotated seedlings was less than in the control. The synthesis rates of δ-aminolevulinic acid and δ-aminolevulinate dehydratase activity in clinorotated seedlings were slower than in the control after 24 h of greening and after 72 h of greening reaching the control values. The activity of Mg-protoporphyrin IX chelatase catalyzing the incorporation of Mg ions in the structure of chlorophyll a, did not change when exposed to clinorotation. The results we obtained show inhibition of the initial stages of chlorophyll biosynthesis in the conditions of simulated microgravity. The light, to a certain extent, decreases the negative effect of microgravity on the formation of the photosynthetic apparatus in plants.     

Effect of horizontal clinorotation on the root system development and on lipid breakdown in rapeseed (Brassica napus) seedlings

Seedlings of Brassica napus were cultivated on a slowly rotating clinostat (1 rpm) or in the vertical control for 5 d. The root growth, the cotyledonary reserves and the transport of 14C-labeled sucrose from cotyledons to root system were studied in both cultural conditions. The biomass (fresh weight) of the root system was 35% higher in the horizontally clinorotated seedlings than in the controls. This increase was correlated with a greater degradation of reserve lipids and faster accumulation of sucrose in the cotyledons. The activity of isocitrate lyase, one of the two enzymes necessary to conversion of lipids into glucids, was also greater in the cotyledons of clinorotated seedlings. The labeling distribution of 14C in the cotyledons, the hypocotyl and the root system after 30, 60 and 120 min of application of 14C-labeled sucrose on the cotyledons showed higher translocation of the cotyledonary sucrose to the root system of clinorotated seedlings. In addition, we studied the effects of clinorotation on the biomass of the excised root system (of 10 d old seedlings) cultivated in a medium containing 1% sucrose. The horizontally clinorotated root system grew more than that of the controls. These results showed that the horizontal clinorotation acted on the root system growth and provoked a higher sucrose translocation from source to sink, i.e. from cotyledons to root system.     

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.     

TNF-alpha-dependent activation of NF-kappa B in human osteoblastic HOS-TE85 cells is repressed in vector-averaged gravity using clinostat rotation

Effects of vector-averaged gravity on tumor necrosis factor (TNF)-alpha-dependent activation of nuclear factor kappa B (NF-kappa B) in human osteoblastic HOS-TE85 cells were investigated by culturing the cells using clinostat rotation (clinorotation). Cell cultures were rotated for 72 h at 40 rpm in a clinostat. At the end of clinorotation, the cells were treated with TNF-alpha for 30 min under stationary conditions. Electrophoretic mobility shift assays revealed that TNF-alpha-dependent activation of NF-kappa B was markedly reduced in the clinorotated cells when compared with the cells in control stationary cultures or after horizontal rotation (motional controls). The NF-kappa B-dependent transactivation was also impaired in the clinorotated cells, as evidenced by a transient transfection assay with a reporter plasmid containing multimerized NF-kappa B sites. Consistent with these findings, the TNF-alpha-dependent induction of endogenous NF-kappa B-responsive genes p105, I kappa B-alpha, and IL-8, was significantly attenuate in clinorotated cells. These results demonstrate that vector-averaged gravity inhibits the responsiveness of osteoblasts to TNF-alpha by repressing NF-kappa B activation.     

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.     

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     

Starch metabolism in germinating soybean cotyledons is sensitive to clinorotation and centrifugation

Soybean (Glycine max [L.] Merr. cv. McCall) seedlings germinated and grew for 6d under the altered gravity conditions of horizontal clinorotation and centrifugation. Both of these conditions resulted in decreased growth relative to the control (vertically rotated) plants. Starch concentration in the cotyledons was lower in the clinorotated plants and was higher in the centrifuged plants compared to the controls. The opposite relationship was noted for total lipid concentration. Of the six starch metabolic enzyme activities measured, only ADP glucose pyrophosphorylase was affected by the gravity treatments; being lower in the cotyledons of the horizontally rotated plants and higher in the cotyledons of the centrifuged plants relative to the control values.     

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.     

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.     

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.     

Superoxide dismutase: an all-purpose gene for agri-biotechnology

The objective of this study was to evaluate the performance of transgenic canola (Brassica napus) plants over-expressing a wheat mitochondrial Mn superoxide dismutase (Mn SOD3.1) subjected to environmental stresses in the field and in controlled environments. Mn SOD3.1 was regulated by either CaMV 35S or Arabidopsis COR78 promoters. RT-PCR and a SOD enzyme activity assays demonstrated Mn SOD3.1 was expressed at both the mRNA and protein levels. Enzyme activity assays exhibited total SOD activity was up to 41.8% and up to 26.7% higher in the 35S:SOD3.1 and in the COR78:SOD3.1 transgenic plants than in the control, respectively. Germination studies, conducted at suboptimal (8°C) and optimal (23°C) temperatures, identified transgenic lines that germinated earlier than the control. In the field under drought conditions, several transgenic lines emerged earlier than the control. In both greenhouse and field environments, several transgenic lines were significantly taller than the control and over 50% of the transgenic canola lines flowered 7–14 days earlier than the control. Over expression of Mn SOD3.1 enhanced heat, drought and cold tolerance both in the field and under artificial stress conditions. This is one of the first tests conducted on transgenic canola plants subjected to field conditions.     

Gravitropic responses of the Avena coleoptile in space and on clinostats. IV. The clinostat as a substitute for space experiments

Gravitropic responses of dark grown oat coleoptiles were measured in weightlessness and under clinorotation on earth. The tests in microgravity were conducted in Spacelab during the IML-1 mission and those on clinostats were conducted in laboratories on earth. The same apparatus was used for both kinds of tests. In both cases autotropism and gravitropic responsiveness were determined. This allowed a quantitative comparison between the plants' responses after receiving the same tropistic stimulations either in weightlessness or on clinostats.
Autotropism was observed with oat coleoptiles responding in weightlessness but it did not occur on clinostats. Gravitropic responsiveness was measured as the ratio between the incremental bending response (degrees curvature) and the corresponding incremental g-dose (stimulus intensity times duration for which it was applied). Plants were tested at either of two stages of coleoptile development (i.e. different coleoptile lengths). From a total of six different kinds of critical comparisons that could be made from our tests that provided data for clinorotated vs weightless plants, three showed no significant difference between responses in simulated vs authentic weightlessness. Three other comparisons showed highly significant differences. Therefore, the validity of clinorotation as a general substitute for space flight was not supported by these results.     

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.     

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.     

The effect of salt stress on lipid peroxidation and antioxidants in the leaf of the cultivated tomato and its wild salt‐tolerant relative Lycopersicon pennellii

The possible involvement of the antioxidative system in the tolerance to salt stress was studied in the cultivated tomato Lycopersicon esculentum Mill. cv. M82 (M82) and its wild salt‐tolerant relative L. pennellii (Corn) D'Arcy accession Atico (Lpa). All analyses, except that of monodehydroascorbate reductase (MDHAR), were performed of the youngest fully‐expanded leaf of control and salt (100 m M NaCl) stressed plants, 4, 7, 10, 14, 18 and 22 days after completing the stress treatment. In Lpa, constitutive level of lipid peroxidation and activities of catalase (CAT) and glutathione reductase (GR) were lower while the activities of superoxide dismutase (SOD), ascorbate peroxidase (APX) and dehydroascorbate reductase (DHAR) were inherently higher than in M82. Relative to M82, lipid peroxidation was much lower and the activities of SOD, CAT and APX were higher in Lpa at 100 m M NaCl. The activity of DHAR decreased more in Lpa than in M82 under salt stress, and the activity of MDHAR, which was lower in Lpa than in M82 under control conditions, increased much more and to a higher level in salt‐treated Lpa plants. GR activity decreased similarly in the two species under salt stress. The results of these analyses suggest that the wild salt‐tolerant Lpa plants are better protected against active oxygen species (AOS), inherently and under salt stress, than the relatively sensitive plants of the cultivated species.     

Aspects of adaptive answering formation in virus-host plant pathosystem for different wheat cultivars under simulating microgravity condition

Investigations of prolonged clinorotation effect on some morphological and physiological parameters under Wheat streak mosaic virus WSMW-infection of Apogee and Lada wheat cultivars were carried out. Experiments were held on universal clinostat CYCLE-2. Clinorotation caused changing of WSMV virions shape and reducing of the virus reproduction. Apogee wheat plants grown under two stress factors (infection and clinorotation) produced more kernels than stationary (motionless) plants, but the average weight of kernel was lower. Under clinorotation changes in host plant-virus system take place and adaptive reactions for simulated microgravity conditions form. These lead to reduction of potyvirus replication.