Seed-Specific Expression of AINTEGUMENTA in Medicago truncatula Led to the Production of Larger Seeds and Improved Seed Germination

The increase of seed size is of great interest in Medicago spp., to improve germination, seedling vigour and, consequently, early forage yield as well as for optimizing seeding techniques and post-seeding management. This study evaluated the effects of the ectopic expression of the AINTEGUMENTA (ANT) cDNA from Arabidopsis thaliana, under the control of the seed-specific USP promoter from Vicia faba, on seed size, germination and seedling growth in barrel medic (Medicago truncatula Gaertn.). All the transgenic T₂ barrel medic lines expressing ANT produced seeds significantly larger than those of control plants. Microscopic analysis on transgenic T₃ mature seeds revealed that cotyledon storage parenchyma cells were significantly larger and contained larger storage vacuoles than those of the untransformed control. Moreover, the percentage of germination was significantly higher and germination was more rapid in transgenic than in control seeds. Our results indicate that the seed-specific expression of ANT in barrel medic led to larger seeds and improved seed germination, and revealed a regulatory role for ANT in controlling seed size development.     

Recalcitrant seeds of horse chestnut lack protein bodies

In recalcitrant seeds of horse chestnut (Aesculus hippocastanum L.), the bulk of protein in axial organs and cotyledons is accounted for by water-soluble proteins (albumins). In the cells of embryo, proteins are predominantly located in the cytosol, whereas the fraction of cell structures precipitate in the range from 1000 to 20000 g, accounting for only an insignificant part of total protein. Among the proteins of this fraction, there were no major components that could play a role of storage proteins. The aim of this work was to study deposition of protein in the vacuoles of cells of recalcitrant seeds of horse chestnut. Light microscopy and specific staining of protein and phytin did not detect protein bodies in the vacuoles of axial organs and cotyledons. Electron microscopy revealed traces of phytin in the vacuoles, but there were no formed globoids or considerable amount of protein therein. It is possible that precisely the absence of typical storage proteins and genetically determined desiccation in the course of maturation of recalcitrant seeds of horse chestnut stipulated preservation of the vacuoles that in mature recalcitrant seeds were not transformed into protein bodies.     

Maintenance of plant cell membrane integrity and function by the immobilisation of protoplasts in alginate matrices

Isolated protoplasts (Vicia faba) suspended in a mannitol solution over a time period of about 10 d undergo a process of aging characterised by increased ethane formation, decrease in oxygen evolution and CO₂ uptake, pigment degradation and a change in certain enzyme activities (ribulose-1,5-bisphosphate carboxylase and protease). The technique of immobilisation of protoplasts in a cross-linked alginate matrix offers a way to delay the appearance of the indicators mentioned. Various possible mechanisms for the increased stability of protoplasts are discussed.     

Effects of the insecticide pyrethroid ii in the ames test,and onhordeum vulgare andvicia faba

The insecticide pyrethroid II, representing synthetic pyrethroids of the second generation, was not found to be mutagenic inSalmonella typhimurium strains TA1535, TA100, TA1538, TA98 and TA97. High concentrations of the tested insecticide inhibited the germination and length of roots of germinating seeds, the height of plants cultivatedin vitro, and slightly increased the frequency of aberrant anaphases and telophase in root-tips ofVicia faba andHordeum vulgare.     

The effect of maleic hydrazide and exogenous DNA on chromosomes ofVicia faba

The mitotic activity of merstematic cells ofVicia faba, the frequency of chromosomal abnormalities and their interchromosomal distribution are evaluated in dependence on the concentration of maleic hydrazide (MH) and on the recovery period. The influence of exogenous DNA of different genetic origin on the course of repair of primary root cells damaged by MH was also studied. Isologous DNA which exhibited a strong repair effect in authors' previous experiments was quite ineffective in the case of maleic hydrazide. Heterologous DNA, on the other hand, had to some extent a parallel effect with MH in breaking down the structural integrity of chromosomes and increased the frequency of aberrations induced by maleic hydrazide in meristematic cells ofVicia faba.     

Chromosome aberrations caused by the effect of ultrasound in the meristematic cells ofvicia faba

In our present work the formation of chromosome aberrations has been studied in dependence on the tima interval between sonication and fixation of the primary root tips of Vicia faba. Maximum occurrence of aberrations was recorded immediately after sonication. The results of our experiments pointed to the fact that the frequency of the induced changes was independent on the sonic waves intensity within the range of 0-2—3-0 W/cm² and on ultrasond treatment duration within the range of 1—20 min. Studies of the distribution of chromosome abnormalities caused by ultrasound between the large and small chromosomes of theVicia faba meristematic cells in various time intervals showed that the frequency of the aberrations in both chromosome groups was proportional to its total metaphase lengths. Analysis of the type of aberrations observed in various time intervals after sonication indicated the simultaneous formation of chromosome and chromatide abnormalities.     

Cytogenetic effect of radiation products of cytosine: Parabanic and oxaluric acids

The two final products of radiolytic degradation of cytosinei.e. parabanic and oxaluric acids were investigated as regards their cytogenetic effects. Broad bean (Vicia faba L.) root meristem was used as experimental material. While the oxaluric acid was not able to induce chromosomal aberrations to a greater extent, the parabanic acid acts as a clastogenic compound. When applied on a resting broad bean root meristem at concentration from 10^-3 to 10^-2 M it induces chromosome and chromatid aberrations, especially breakages. Their frequency reaches 9–12% at the highest tested concentration. The same concentrations of the parabanic acid increases mitotic index of the first cell generation of primary root.     

Participation of exogenous DNA in the repair processes of meristematicVicia faba cells injured by monofunctional alkylating agens ethylmethane sulphonate

The effect of exogenous DNA of syngeneic origin on the course of reparation of meristematic cells ofVicia faba primary roots followed after treatment with monofunctional alkylating agent, ethyl methanesulphonate (EMS), was tested. Time course of alternations in mitotic activity of investigated cellular population and the dynamics of formation of postmetaphase chromosomal aberrations was evaluated. A reparation of damaged cells was significantly supported by syngeneic DNA; its application induced an increased incidence of cellular division already in the early intervals of the repair which was accompanied by concomitant decrease of microscopically detectable rupture in the chromosomes. The study performed on the localization of induced damages occurring in metaphase chromosomes pointed out an increased sensitivity of small chromosomes ofVicia faba to EMS. Similarly, a reparative action of syngeneic DNA was exhibited by significant decrease of aberrations frequency, predominantly in the same chromosomal group. Per cent representation of individual types, not affected by the action of syngeneic DNA, was established by detailed classification of induced aberrations. In both cases, isochromatide breaks were found of greatest predominance.     

Contribution to the study on the reparative effect of exogenous DNA in the irradiated meristem ofVicia faba

The present report is focused on the study of participation of exogenous DNA in the process of postirradiation reparation of meristematic cells ofVicia faba primary roots. It is aimed at comparison of the positive reparative effect of isologous DNA with postirradiation action of heterologous DNA in its native, thermally denatured and DNAase-degraded forms, or DNA degraded by ultrasound, and with the effect of other biologically important macromolecules (RNA, histone, heparin, and dextran sulphate). For this purpose, the roots ofVicia faba irradiated by 150 r exposure were cultivated in solutions containing the above substances for an appropriate time interval. In squash slides both mitotic activity of the investigated cell population and frequency of postmetaphase chromosomal aberrations induced by radiation were evaluated. It was shown that a stimulation of cell division and reparation of chromosome damages were supported exclusively by isologous DNA. On the contrary, exogenously applied heterologous DNA increased postirradiation frequency of aberrations; maintenance of native structure of applied DNA was an essential condition for the above effect. Other macromolecules investigated on the course of postirradiation reparation ofVicia faba meristematic cells were without effect.     

Cytotaxonomic study of theVicia cracca complex 1. Czechoslovak taxa

Three races ofVicia cracca L. have been found on the territory of Czechoslovakia, two with the chromosome number 2n=14 and one with 2n=28. Diploid races seem to be more primitive and are less widely distributed. They occur mostly in the primary communities while the tetraploid race is very plastic in the ecological respect and is common both in primary and secondary communities. These races are characterized by some morphological and ecological features. As far as the vertical distribution is concerned, one of the diploid races and the tetraploid occur mostly in the lowland and in the colline zone, the second diploid race has a mountain character. The related tetraploid speciesVicia oreophila ?ertová, with the chromosome number 2n=28 is distributed at similar altitudes.     

Histogenesis and localization of non-specific esterase in root tip

A procedure was developed for satisfactory freeze-sectioning of root tips. The use of Ca formol-fixed material kept and frozen in Holt's syrup is recommended. The existence and different localization of 2 fractions of non—specific esterase was verified in root tips ofVicia faba. The same results were revealed in fixed and unfixed material. The dynamics ofin situ reaction was followed with respect to optimal incubation time. The results with substrates of different chain length support the existence of 2 fraction of the studied enzyme, none of which, concerning substrate specificity, is a lipase. It follows from the present studies inVicia faba and other species (Cucurbita pepo, Lupinus albus, Pisum sativum Zea mays), that non-specific esterase localization is not directly given by histogenesis.     

Comparative effects of colchicine,caffeine

The effect of colchicine, caffeine and hydroquinone on nodal meristems ofCallisia fragrans has been studied. The polyploidy have been induced following 4 and 8 hours of treatment in 0.5% colchicine. The persistence of polyploidy in emerging roots even after 60 d. of recovery in the soil indicates that the colchicine affected not only the nodal roots but also the nodal tissue of the plant. The occurence of both diploid and polyploid roots in the same node has been attributed to the differential penetration of the compound to the different zones of tissue. The increase in the division frequency following certain period of recovery was also observed. Caffeine induced only different subnarcotic effects. The formation of the binucleate cells, (which is very common inVicia faba) has been found to be very rare in the present material. No significant results have been obtained following the hydroquinone treatment excepting some common subnarcotic effects.     

Membrane anchors effectively traffic recombinant human glucocerebrosidase to the protein storage vacuole of Arabidopsis seeds but do not adequately control N-glycan maturation

Key message

Human glucocerebrosidase with vacuolar anchoring domains was targeted to protein storage vacuoles (PSVs) of Arabidopsis seeds, but unexpectedly via the Golgi complex. PSV-targeting to effectively avoid problematic N-glycans is protein dependent.

Abstract

Plant-specific N-glycosylation patterns elaborated within the Golgi complex are a major limitation of using plants to produce biopharmaceuticals as the presence of β1,2 xylose and/or α1,3 fucose residues on the recombinant glycoprotein can render the product immunogenic if administrated parenterally. A reporter protein fused to a vacuolar membrane targeting motif comprised of the BP-80 transmembrane domain (TMD), and the cytoplasmic tail (CT) of α-tonoplast intrinsic protein (α-TIP) is delivered to protein storage vacuoles (PSVs) of tobacco seeds by ER-derived transport vesicles that bypass the Golgi complex. This prompted us to investigate whether a pharmaceutical glycoprotein is targeted to PSVs using the same targeting sequences, thus avoiding the unwanted plant-Golgi-specific complex N-glycan modifications. The human lysosomal acid β-glucosidase (glucocerebrosidase; GCase) (EC 3.2.1.45) fused to the BP-80 TMD and α-TIP CT was produced in Arabidopsis thaliana wild-type (Col-0) seeds. The chimeric GCase became localized in PSVs but transited through the Golgi complex, as indicated by biochemical analyses of the recombinant protein’s N-glycans. Our findings suggest that use of this PSV-targeting strategy to avoid problematic N-glycan maturation on recombinant therapeutic proteins is not consistently effective, as it is likely protein- and/or species-specific.     

Effects of salt stress on nitrogenase activity and growth of four legumes

The effects of salt stress on nitrogenase (N2-ase) activity, growth and nitrogen content ofVicia faba (L.),Medicago sativa (L.) Merrill,Glycine max andVigna sinensis (L.) were investigated. Four levels of salinity were applied and salt treatments were imposed on inoculated and N-fertilized plants.M. sativa tolerated mild levels of salinity but higher salt concentrations depressed N₂-ase activity of this species. The other three legumes were considerably affected by salt treatments, and N₂-ase activity was significantly reduced by salinity. Vicia faba, carrying elongate nodules, could restore a partial N₂-ase activity upon recovery from salt stress whereasG. max andV. sinesis, both with spherical nodules, could not regain significant activity when salinity was removed. Salt stress retarded growth of both inoculated and N-fertilized plants. The nitrogen content of both treatments was also affected by salinity and the effect was more severe for inoculated than N-fertilized plants.     

Ethylene-enhanced ethylene oxidation inVicia faba

The alkene oxygenase (AO) of fababean (Vicia faba L.) converts ethylene to ethylene oxide. Treatment of fababeans with 10μl/liter ethylene increases the activity of this enzyme within 2 hours of ethylene treatment. Though other alkenes were taken up by fababean seedlings, ethylene was the most active in inducing AO activity. The ability of ethylene to increase AO was blocked 60% by cycloheximide, an inhibitor of protein synthesis, and 35% by AgNO₃, an inhibitor of ethylene action.     

Woloszynskia stoschii andGymnodinium indicum,two new freshwater dinoflagellates from India: Morphology,reproduction and cytology

The distinctive morphological and karyological features of the two new species, collected from freshwater habitats in North India, are presented and compared with those of related taxa of the generaWoloszynskia andGymnodinium. Asexual reproduction takes place through zoospore formation and sexual reproduction through planozygotes inWoloszynskia stoschii.     

Production of plant growth substances by rhizosphere mycoflora of broad bean and cotton

The culture filtrates of the rhizosphere fungi of broad bean (Vicia faba L.) and cotton (Gossypium barbadense L.) were analysed for the presence of plant growth substances of auxin and gibberellins nature. Bioassay test and chromatographic analysis indicated that these fungi, each synthesized different auxins in their culture medium. These auxins were indole compounds. Similarly the rhizosphere fungi produced in their culture medium some gibberellins and gibberellin-like substances.     

Ultrastructural development of plastids in cherry peppers during fruit ripening

The fine structure of plastids in the fruit of cherry peppers was studied during the various stages of ripening. The color change of fruit during ripening is due to the quantitative change of such pigment components as chlorophyll, carotenoid and anthocyanin. Plastid metamorphosis takes place in relationship to the disappearance of chlorophyll and the new formation of carotenoids. The membrane system of plastids degenerates through ripening, although a little differentiation is observed in young plastids of creamy fruits. In parallel wity the color change of fruit from cream to orange, the osmiophilic globules increase in both number and size. As ripening proceeds further, the large osmiophilic globules seem to be gradually transformed into the needle shaped crystalloids of carotenoid pigments which are the remarkable feature of the chromoplasts in red-ripe fruit. The relationship between the development of chromoplasts and the increase and decrease of some pigments is also discussed.     

Studies on the rhizosphere mycoflora of broad bean and cotton. Effect of fungal filtrate on plant growth

The dominating rhizosphere fungi of broad bean (Vicia faba Linn.) variety “Giza 1”, and cotton (Gossypium barbadense Linn.) variety “Giza 47”, were grown in liquid medium. After 10 days, filtrates were obtained and sterilized by filtration through sintered-glass filter. Plants were grown in sterile sand which was supplemented with nutrient solution. Every plant was irrigated with fungal filtrate, unconsumed medium, and water. The filtrates of the rhizosphere fungi of both broad bean and cotton stimulated plant growth.     

Cytogenetic studies on the effect of chronic gamma irradiation onVicia faba

Vicia faba seeds (cv. Giza 1) were planted in the Inshas gamma radiation field where they were chronically irradiated during the whole life of the plant. The percentage of the induced abnormal P.M.Cs, as well as the frequency of abnormal P.M.Cs in the different meiotic stages were proportional with the given doses. The main types of chromosome aberrations were anaphase and telophase bridges, fragmentation and lagging chromosomes. The nearest plants to the source showed an inhibition of shoot growth, flower and seed sterility and irregular branching. At the dosage levels used irradiation had no effect on pollen fertility. Seeds of the 1st filial generation were used for both mitotic and meiotic studies. The percentage of the mitotic abnormalities was proportional with the doses. The most dominant type of anomaly was the presence of micronuclei in the different stages of mitosis and in the resting cells. Irradiation affected also other types of anomaliese.g. lagging chromosomes, fragments, bridges...etc. Meiosis, and pollen fertility (2nd generation) were normal.