Can arsenic-phytochelatin complex formation be used as an indicator for toxicity in Helianthus annuus?

The formation of arsenic-phytochelatin (As-PC) complexes is thought to be part of the plant detoxification strategy for arsenic. This work examines (i) the arsenic (As) concentration-dependent formation of As-PC complex formation and (ii) redistribution and metabolism of As after arrested As uptake in Helianthus annuus. HPLC with parallel ICP-MS/ES-MS detection was used to identify and quantify the species present in plant extracts exposed to arsenate (As(V)) (between 0 and 66.7 micromol As l-1 for 24 h). At As concentrations below the EC50 value for root growth (22 micromol As l-1) As uptake is exponential, but it is reduced at concentrations above. Translocation between root and shoot seemed to be limited to the uptake phase of arsenic. No redistribution of As between root and shoot was observed after arresting As exposure. The formation of As-PC complexes was concentration-dependent. The amount and number of As-PC complexes increased exponentially with concentration up to 13.7 micromol As l-1. As(III)-PC3 and GS-As(III)-PC2 complexes were the dominant species in all samples. The ratio of PC-bound As to unbound As increased up to 1.3 micromol As l-1 and decreased at higher concentrations. Methylation of inorganic As was only a minor pathway in H. annuus with about 1% As methylated over a 32 d period. The concentration dependence of As-PC complex formation, amount of unbound reduced and oxidized PC2, and the relative uptake rate showed that As starts to influence the cellular metabolism of H. annuus negatively at As concentrations well below the EC50 value determined by more traditional means. Generally, As-PC complexes and PC-synthesis rate seem to be the more sensitive parameters to be studied when As toxicity values are to be estimated.     

Organogenesis and embryogenesis inHelianthus tuberosus and in the interspecific hybridHelianthus annuus ×Helianthus tuberosus

A method of plant regeneration from cotyledons ofHelianthus tuberosus, Helianthus annuus ×Helianthus tuberosus and for the backcross of the interspecific hybrids onH. annuus was developed. Induction of somatic embryogenesis and plantlet regeneration from anther culture of the interspecific hybridsH. annuus ×H. tuberosus is reported.Cotyledons were cultured on Murashige and Skoog basal medium (MS) supplemented with indole-3-acetic acid (IAA) and 6-furfurylaminopurine (kinetin) or N⁶-benzylaminopurine (BAP). Shoot regeneration occurred on most of the media tested, but the best results were obtained on media with a high concentration of cytokinins (BAP or kinetin: 4 mg l^–1) and lower concentration of auxin (IAA: 0.5–1 mg l^–1).Embryogenic callus and adventitious buds were initiated from only two anthers of the hybridH. annuus ×H. tuberosus cultured on the MS medium containing BAP (0.2 mg l^–1) and 1-naphtalenacetic acid (NAA: 0.1 mg l^–1). Prolonged culture of these embryogenic calli and buds on the original medium with successive subculture on MS basal medium without growth regulators resulted in embryo formation and shoot differentiation. The plantlets, after rooting, were established in soil.     

Untersuchungen zur Funktionsänderung der Microbodies in den Keimblättern von Helianthus annuus L.

Summary The enzyme patterns in sunflower cotyledons indicate that the glyoxysomal function of microbodies is replaced by the peroxisomal function of these organelles during the transition from fat degradation to photosynthesis. The separation of the microbody population into glyoxysomes and peroxisomes during this transition period is reported. The mean difference in density between the activity peaks of glyoxysomal and peroxisomal marker enzymes on a sucrose gradient was calculated to be 0.007±0.004 g/cm³ and turned out to be significant (t=7.8>4.04=t _5;0.01). The activity peak of catalase coincides with that of isocitrate lyase in early stages of development, but shifts to the activity peak of peroxisomal marker enzymes during the transition period. No isozymes of the catalase could be detected by gel electrophoresis in the microbodies with the two different functions.During the rise of the peroxisomal marker enzymes no synthesis of the common microbody marker, catalase, could be demonstrated using the inhibitor allylisopropylacetamide. Using D₂) for density labeling of newly-formed catalase, no difference is observed between the density of catalase from cotyledons grown on 99.8% D₂O during the transition period and the density of enzyme from cotyledons grown on H₂O. The activity of particulate glycolate oxidase is reduced 30–50% by allylisopropylacetamide, but is not affected by D₂O. The chlorophyll formation in the cotyledons is strongly inhibited by both substances.     

Auxin — Calcium Interaction in Adventitious Root Formation in the Hypocotyl Explants of Sunflower (Helianthus annuus L.)

Auxin-calcium interaction has been studied to understand their involvement in adventitious root initiation from the hypocotyl explants of sunflower (Helianthus annuus L.). When hypocotyl explants were cultured on MS medium (containing calcium), 1 mg l^-1 IAA was found to be optimal for root induction. However, the hypocotyl explants washed in EGTA (10_-5M) solution for the removal of extracellular calcium, when cultured on medium containing IAA and calcium, exhibited enhanced rooting response. When EGTA-washed explants were cultured on the medium supplemented with lanthanum chloride (10^-6 and 10^-5M), it resulted in the inhibition of the rooting response and this inhibitory effect could be alleviated by the simultaneous addition of IAA. Similar observations have been made by using calcium channel blockers, verapamil and TMB-8, and also a calmodulin inhibitor, trifluoperazine. A net influx of extracellular calcium in the differentiating cells is thus presumed to accompany the auxin-induced response. These results have been discussed in light of initial lack of polarity in the decapitated hypocotyl segments subjected to auxin treatment.     

Cytotoxic constituents from the leaves of Jerusalem artichoke (Helianthus tuberosus L.) and their structure–activity relationships

Bioassay-directed phytochemical study of the Jerusalem artichoke (Helianthus tuberosus L.) leaves led to the isolation of a new sesquiterpene lactone of 3-Hydroxy-8β-tigloyloxy-1,10-dehydroariglovin (1), ten known sesquiterpene lactones (2–11) and two known flavones (12–13). Their chemical structures were elucidated on the basis of NMR (1D and 2D) and mass spectroscopic analysis. The cytotoxic activities of those compounds were subsequently tested against the MCF-7, A549 and HeLa cancer cells lines. The results indicated that sesquiterpene lactones 1–11 exhibited consistent cytotoxicity against all three cancer cell lines, while flavones 12 and 13 showed selective inhibitory activity against HeLa cell lines. Among them, compound 3 exhibited significant growth inhibitory activity against all three cell lines. The IC₅₀ values of compound 3 against MCF-7, A549 and HeLa were 1.97 ± 0.04, 7.79 ± 0.44, 9.87 ± 0.20 μg/ml, respectively. In addition, some structure–activity relationships of these sesquiterpene lactones for cytotoxicity were explored and summarized in this study.     

Cytogenetic analysis of interspecific hybrid Helianthus praecox × H. annuus, its parental forms, and two backcrosses

Meiosis of wild (Helianthus praecox) and cultivated (H. annuus) sunflower species, their F₁ interspecific hybrid, and two backcross generations were studied. A low level of meiotic chromosomal aberrations in the parents was detected. Interspecific hybridization between wild and cultivated sunflower accessions led to a considerable (50%) increase in the rate of meiotic aberrations in microsporocytes. Backcrossing of hybrids to cultivated sunflower decreased the chromosomal aberration rate to 12.8% in BC₁ and to 9.6% in BC₂. Cytological stability of plants was restored only in BC₂.     

Induction of a photomixotrophic plant cell culture of Helianthus annuus and optimization of culture conditions for improved α-tocopherol production

Tocopherols, collectively known as vitamin E, are lipophilic antioxidants, which are synthesized only by photosynthetic organisms. Due to their enormous potential to protect cells from oxidative damage, tocopherols are used, e.g., as nutraceuticals and additives in pharmaceuticals. The most biologically active form of vitamin E is α-tocopherol. Most tocopherols are currently produced via chemical synthesis. Nevertheless, this always results in a racemic mixture of different and less effective stereoisomers because the natural isomer has the highest biological activity. Therefore, tocopherols synthesized in natural sources are preferred for medical purposes. The annual sunflower (Helianthus annuus L.) is a well-known source for α-tocopherol. Within the presented work, sunflower callus and suspension cultures were established growing under photomixotrophic conditions to enhance α-tocopherol yield. The most efficient callus induction was achieved with sunflower stems cultivated on solid Murashige and Skoog medium supplemented with 30 g l^?1 sucrose, 0.5 mg l^?1 of the auxin 1-naphthalene acetic acid, and 0.5 mg l^?1 of the cytokinin 6-benzylaminopurine. Photomixotrophic sunflower suspension cultures were induced by transferring previously established callus into liquid medium. The effects of light intensity, sugar concentration, and culture age on growth rate and α-tocopherol synthesis rate were characterized. A considerable increase (max. 230 %) of α-tocopherol production in the cells was obtained within the photomixotrophic cell culture compared to a heterotrophic cell culture. These results will be useful for improving α-tocopherol yields of plant in vitro cultures.     

Chemical role of α-tocopherol in salt stress mitigation by improvement in morpho-physiological attributes of sunflower (Helianthus annuus L.)

Elevated concentrations of salts in soil and water represent abiotic stresses. It considerably restricts plant productivity. However, the use of alpha-tocopherol (α-toc) as foliar can overcome this problem. It can improve crop productivity grown under salinity stress. Limited literature is documented regarding its optimum foliar application on sunflower. That’s why the need for the time is to optimize α-toc foliar application rates for sunflower cultivated in salt-affected soil. A pot experiment was performed to select a better α-toc foliar application for mitigation of salt stress in different sunflower cultivars FH (572 and 621). There were 2 levels of salts, i.e., control (no salt stress) and sodium chloride (120 mM) and four α-toc foliar application (0, 100, 200, and 300 mg L^?1). Results showed that foliar application of 100 mg/L- α-toc triggered the remarkable increase in fresh shoot weight, fresh root weight, shoot, and root lengths under salinity stress in FH-572 and FH-621 over 0 mg/L- α-toc. Foliar application of 200 mg/L- α-toc was most effective for improvement in chlorophyll a, chlorophyll b, total chlorophyll and carotenoids compared to 0 mg/L- α-toc. Furthermore, an increase in A was noted in FH-572 (17%) and FH-621 (22%) with α-toc (300 mg L^?1) application under saline condition. In conclusion, the 100 and 200 mg/L- α-toc are the best application rates for the improvement in sunflower FH-572 and FH-621 growth, chlorophyll contents and gas exchange attributes. Further investigations are needed to select a better foliar application rate between 100 and 200 mg/L- α-toc at the field level under the different agro-climatic zone and soil types.     

Is sunflower (Helianthus annuus) at risk to damage from Ophraella communa,a natural enemy of common ragweed (Ambrosia artemisiifolia)?

Ophraella communa is being studied for augmentative releases in eastern Canada for Ambrosia artemisiifolia (common ragweed) control. Host specificity studies with cohorts of insects and a life table approach with individual insects were utilized to evaluate the risk of damage to sunflower. Predicting relative non-target attack and when and where this may occur is based on relative acceptability, suitability, and availability of host and non-host plants. We conclude the risk of O. communa adult-stage beetles attacking sunflower plants is negligible. In 20% open cages, over 98% of the adult O. communa left sunflower plants with few or no eggs laid. When the preferred host plants (A. artemisiifolia) are completely defoliated, the first instar larvae move to nearby sunflower plants in the immediate vicinity (up to 20-30 cm) and begin foraging. O. communa larvae can significantly damage sunflower, but 50% of the first instar larvae will die feeding on sunflower plants. The effect of low oviposition and high mortality will result in no increase in the O. communa population. Within 30 days, the O. communa population increased 208 times when feeding on ragweed and decreased 4.2 times when feeding on sunflower. Newly emerged adults leave the sunflower plants in search of ragweed. Damage to sunflower could occur during augmentative releases of O. communa, but this would only occur when a significantly large, advancing insect population has completely defoliated A. artemisiifolia within or adjacent to a sunflower field.     

Uptake and distribution of zinc,cadmium, lead and copper in Brassica napus var. oleífera and Helianthus annus grown in contaminated soils

Brassica napus var. oleifera and Helianthus annus were grown in artificially contaminated soils. Accumulation and translocation of the environmental pollutants zinc, cadmium, lead, and copper, was evaluated in different portions of the plants at two harvesting times. The distribution into the plants of these metal ions, as well as their capacity for contaminant phytoextraction and accumulation was assessed. For this purpose, an analytical method utilizing focused ultrasound employed for extraction and stripping voltammetry for measurement has been optimized and validated for the simultaneous measurement of Zn, Cd, Pb, and Cu in plant extracts.     

Comparative Study of the Rhizospheric Microflora of Sunflower Cultivars of Helianthus annuus (Asteraceae,Magnoliópsida) Grown on Soils with Anthropogenic Polyelemental Anomalies

Biology Bulletin - In a laboratory pot experiment, two mutant cultivars of sunflower (Helianthus annuus cv. r2p2 and Helianthus annuus cv. r5n1) were grown on soils with anthropogenic polyelemental...     

Formation of zeatin allylic phosphate by the microsomal fractions of bulb disks of Iris × hollandica Tub. and tubers of Helianthus tuberosus L.

It is shown that in bulbous Iris zeatin originates from a nucleotide. This nucleotide is probably zeatin-allylic-phosphate, in which a phosphate group is attached to the isoprenoid side-chain of zeatin. The formation of zeatin-allylic-phosphate from t-zeatin and 8-[¹⁴C]-zeatin by the microsomal fractions of Iris bulb disks and Helianthus tubers was demonstrated. The responsible enzyme was partially purified. 5-AMP was found to be a phosphate group delivering substrate. Adenosine and adenine inhibited the enzyme reaction. The significance of the results is discussed in relation to cytokinin biosynthesis and the occurrence of bud blast in Iris.     

Assay for doubled haploid sunflower (Helianthus annuus) plant production by androgenesis: fact or artifact? Part 1. In vitro anther culture

Sunflower anthers placed on solid medium developed calli and embryos after 12 days. Embryogenesis was improved by the addition of 0.1% polyvinylpyrrolidone (PVP) that alleviated anther and medium browning. As in other species, genotypic variability was an important parameter in the anther response and a medium genotype interaction was suggested with a different PVP effect depending on the genotype. Embryo germination was largely increased by the successive use of germination media with decreasing sucrose concentrations (10%6%3%). Histological examination of the anthers during the first ten days of culture showed that, under our conditions, the embryos were of somatic origin, arising directly from the anther wall on the outside or inside of the anther loculus, or indirectly from proliferating anther wall- or connective tissue-derived callus. Finally, the ploidy status of 78 embryo-derived plants was determined by Feulgen stain or flow cytometry: all plants were diploid (2n=34).Abbreviations PVP polyvinylpyrrolidone     

Sphingolipid base modifying enzymes in sunflower (Helianthus annuus): cloning and characterization of a C4-hydroxylase gene and a new paralogous Δ8-desaturase gene

Sphingolipids are components of plant cell membranes that participate in the regulation of important physiological processes. Unlike their animal counterparts, plant sphingolipids are characterized by high levels of base C4-hydroxylation. Moreover, desaturation at the Δ8 position predominates over the Δ4 desaturation typically found in animal sphingolipids. These modifications are due to the action of C4-hydroxylases and Δ8-long chain base desaturases, and they are important for complex sphingolipids finally becoming functional. The long chain bases of sunflower sphingolipids have high levels of hydroxylated and unsaturated moieties. Here, a C4-long chain base hydroxylase was functionally characterized in sunflower plant, an enzyme that could complement the sur2Δ mutation when heterologously expressed in this yeast mutant deficient in hydroxylation. This hydroxylase was ubiquitously expressed in sunflower, with the highest levels found in the developing cotyledons. In addition, we identified a new Δ8-long base chain desaturase gene that displays strong homology to a previously reported desaturase gene. This desaturase was also expressed in yeast and was able to change the long chain base composition of the transformed host. We studied the expression of this desaturase and compared it with that of the other isoform described in sunflower. The desaturase form studied in this paper displayed higher expression levels in developing seeds.     

In vitro morphogenesis of sunflower (<Emphasis Type="Italic">Helianthus annuus</Emphasis>) hypocotyl protoplasts: the effects of protoplast density,haemoglobin and spermidine

Sunflower, as one of the most important oil-producing crops, represents an important target for genetic improvement through gene transfer or somatic hybridization. Unfortunately, sunflower is recognized as recalcitrant to in vitro culture. The aim of our paper was to improve sunflower protoplast regeneration. Three cultivars (Romanian hybrids) and one inbred line were used for protoplast isolation from etiolated hypocotyls. Isolated protoplasts were embedded in alginate disks and cultured in two plating densities, using two culture regimes as indicated by previous authors. Plating efficiency, callus development and plant regeneration were evaluated as well as old callus histology. In cv. ‘Select’, the effects of 1:50 haemoglobin and 1 mM spermidine were assayed on asymmetric division and/or plating efficiency. Plant regeneration from hypocotyl protoplasts was achieved for two cvs., ‘Florom 328’ and ‘Turbo’, with the former proving once more its totipotency. The best culture regime proved to be as recommended by Krasnyanski and Menczel (1993), but the best density in the culture medium was the highest ever tested, 8 × 10⁵ pp ml⁻¹. Moreover, the histology of old green compact protoplast-derived callus revealed a very well organized structure suggesting senescence. In the non-responsive cv. ‘Select’, haemoglobin was found to stimulate protoplast asymmetric division and the development of heart-shaped embryo-like structures, while spermidine stimulated overall protoplast plating efficiency.     

Source of Ca,Cd, Cu,Fe, K,Mg, Mn,Mo and Zn in grains of sunflower (Helianthus annuus) grown in nutrient solution: root uptake or remobilization from vegetative organs?

Plant and Soil - This study investigated the possible source organs delivering several trace elements to seeds (root uptake versus net remobilization), by studying changes in biomass and element...     

Effect of dichlorophenolindophenol, dichlorophenolindophenol-sulfonate, and cytochromec on redox capacity and simultaneous net H+/K+ fluxes in aeroponically grown seedling roots of sunflower (Helianthus annuus L.): New evidence for a plasma membrane CN−-resistant redox chain

Summary Excised roots from axenically grown sunflower seedlings reduced or oxidized exogenously added 2,6-dichlorophenolindophenol (DCIP), DCIP-sulfonate (DCIP-S), and cytochromec, and affected simultaneous H⁺/K⁺ net fluxes. Experiments were performed with nonpretreated living and CN^–-pretreated poisoned roots (control and CN^–-roots). CN^–-roots showed no H⁺/K⁺ net flux activity but still affected the redox state of the compounds tested. The hydrophobic electron acceptor DCIP decreased the rate of H⁺ efflux in control roots with extension of the maximum rate and optimal pH ranges, then the total net H⁺ efflux (H⁺) equalled that of the roots without DCIP. The simultaneously measured K⁺ influx rate was first inhibited, then inverted into efflux, and finally influx recovered to low rates. This effect could not be due to uptake of the negatively charged DCIP, but due to the lower H⁺ efflux and the transmembrane electron efflux caused by DCIP, which would depolarize the membrane and open outward K⁺ channels. The different H⁺ efflux kinetics characteristics, together with the small but significant DCIP reduction by CN^–-roots were taken as evidence that an alternative CN^–-resistant redox chain in the plasma membrane was involved in DCIP reduction. The hydrophilic electron acceptor DCIP-S enhanced both H⁺ and K⁺ flux rates by control roots. DCIP-S was not reduced, but slightly oxidized by control roots, after a lag, while CN^–-roots did not significantly oxidize or reduce DCIP-S. Perhaps the hydrophobic DCIP could have access to and drain electrons from an intermediate carrier deep inside the membrane, to which the hydrophilic DCIP-S could not penetrate. Also cytochromec enhanced H⁺ and K⁺, consistent with the involvement of the CN^–-resistant redox chain. Control roots did not reduce but oxidize cytochromec after a 15 min lag, and CN^–-roots doubled the rate of cytochromec oxidation without any lag. NADH in the medium spontaneously reduced cytochromec, but control or CN^–-roots oxidized cytochromec, despite of the presence of NADH. In this case CN^–-roots were less efficient, while control roots doubled the rate of cytochromec oxidation by CN^–-roots, after a 10 min lag in which cytochromec was reduced at the same rate as the medium plus NADH did. CN^–-roots seemed to have a fully activated CN^–-resistant branch. The described effects on K⁺ flux were consistent with the current hypothesis that redox compounds changed the electric membrane potential (de- or hyperpolarization), which induces the opening of voltage-gated in- or outward K⁺ channels.Abbreviations Cyt c cytochromec - DCIP 2,6-dichlorophenolindophenol - DCIP-S 2,6-dichlorophenolindophenol 3-sulfonate - HCF(III) hexacyanoferrate (III) - PM plasma membrane - SHAM salicylhydroxamic acid - VH⁺ and VK⁺ H⁺ efflux and K⁺ influx rates - H⁺ and K⁺ total H⁺ efflux and K⁺ influx at the end of the experiment - H⁺ and K⁺ buffering power of the titrated medium