The potential of production of sulfated polysaccharides from Porphyridium

Summary The environmental conditions prevailing in Israel make marine algae an attractive crop for the production of valuable chemicals. A marine species of Porphyridium seems to fit this purpose.The unicellular red alga Porphyridium is encapsulated by a polysaccharide envelope that is present in the gel state. This polysaccharide is an acidic heteropolymer composed of sulfated sugars. It forms ionic bridges through divalent cations, thus reaching a very high molecular weight. The thickness of the polysaccharide capsule varies according to the phase of growth and the growth conditions. Its outer part dissolves in the growth medium, which becomes progressively more viscous. Sulfated polysaccharides form theramlly reversible gels similar to agar and carrageenan, which are usually extracted from marine macroalgae. These gels have been finding increasing use in commercial applications as gelling agents, thickeners, stabilizers, and emulsifiers.We have done experiments on the cultivation of a marine species of Porphyridium for the production of polysaccharides. This unicellular alga has an advantage over the macroalgae due to its relatively faster growth rate and the possibility to regulate its growth. The potential for production of the polysaccharide, both that dissolved in the external medium and that attached to the cell (including an intracellular fraction), and the effects of growth conditions on productivity were suudied in the laboratory. Porphyridium was also cultivated outdoors in seawater in 1-m² ponds and its growth potential investigated.     

Mechanics of circadian pulvini movements in Phaseolus coccineus L.

The circadian movement of the lamina of primary leaves of Phaseolus coccineus L. is mediated by antagonistic changes in the length of the extensor and flexor cells of the laminar pulvinus. The cortex of the pulvinus is a concentric structure composed of hexagonal disc-like cells, arranged in longitudinal rows around the central stele. Observations with polarization optics indicate that the cellulose microfibrils are oriented in a hoop-like fashion in the longitudinal walls of the motor cells. This micellation is the structural basis of the anisotropic properties of the cells: tangential sections of the extensor and flexor placed in hypotonic mannitol solutions showed changes only in length. As a consequence a linear correlation between length and volume was found in these sections. Based on the relationship between the water potential (which is changed by different concentrations of mannitol) and the relative volume of the sections and on the osmotic pressure at 50% incipient plasmolysis, osmotic diagrams were constructed for extensor and flexor tissues (cut during night position of the pulvinus). The bulk moduli of extensibility, , were estimated from these diagrams. Under physiological conditions the values were rather low (in extensor tissue below 10 bar, in flexor tissue between 10 to 15 bar), indicating a high extensibility of the longitudinal walls of the motor cells. They are strongly dependent on the turgor pressure at the limits of the physiological pressure range.In well-watered plants, the water potentials of the extensor and flexor tissues were surprisingly low,-12 bar and-8 bar, respectively. This means that the cells in situ are by no means fully turgid. On the contrary, the cell volume in situ is similar to the volume at the point of incipient plasmolysis: the cell volumes of extensor and flexor cells in situ were only 1.01 times and 1.1 times larger, respectively, than at the point of incipient plasmolysis, whereas at full turgidity (cells in water) the corresponding factors were 1.8 and 1.5. It is suggested that the high elasticity of the longitudinal walls, the anisotropy of the cell walls, and the low water potential of the sections which is correlated with slightly stretched cell walls in situ, are favourable and effective for converting osmotic work in changes in length of the pulvinus cells, and thus for the up and down movement of the leaf.Symbols volumetric elastic modulus - _i instantaneous volumetric elastic modulus - _i stationary volumetric elastic modulus - weight-averaged stationary bulk modulus of extensibility - ₀ osmotic pressure of the vacuole of a cell at the point of incipient plasmolysis - weight-averaged osmotic pressure of the vacuoles of the tissue at 50% incipient plasmolysis - water potential     

Expression of A and B Types of Monoamine Oxidase in Differentiated Neuroblastoma Hybrid Cells

The total activities of monoamine oxidase (MAO) and the ratio of type B/type A activities were determined in mouse neuroblastoma N1E-115 cells, and in NX31T and NG108-15 hybrid cells derived from mouse neuroblastoma X rat sympathetic ganglion hybrid or mouse neuroblastoma X rat glioma hybrid cells. N1E-115 and NX31T cells possessed type A activities exclusively, although NG108-15 cells showed both type A (65-90%) and type B (10-35%) MAO activities. The activity of type A MAO in NX31T and N1E-115 cells was relatively constant during culturing periods in the presence or absence of dibutyryl cyclic AMP (Bt2cAMP), whereas total MAO activity and the ratio of type B MAO/type A MAO in NG108-15 cells increased as a function of culture periods. Prostaglandin E1 (PGE1) and theophylline, the best known combination to increase intracellular cyclic AMP content of NG108-15 cells, caused similar increases of MAO and of the type B/type A ratio in NG108-15 cells. The results suggest that MAO activity and expression of MAO B activity are regulated in NG108-15 cells in a cyclic AMP-dependent manner.     

Improved preservation of the form and contents of wall vesicles and the golgi apparatus in freeze substituted hyphae ofSaprolegnia

Summary Secretory vesicles involved in cell wall synthesis (wall vesicles) and the Golgi apparatus have been compared in conventionally fixed and freeze substituted hyphae of the oomycete fungusSaprolegnia ferax. Wall vesicles freeze substituted in various fluids range from spherical to tubular and contain an intensely staining, phosphorous rich matrix. In contrast diverse conventional fixations cause artefactual constrictions in most tubular vesicles and loss of their intensely staining contents. These data are interpreted to show the existence of an intravesicular skeletal system, with cellular regulation, to determine vesicle morphology and intravesicular synthesis of a hypothetical phosphorylated glycolipid cell wall precursor. Whilst freeze substitution gives superior preservation of wall vesicle morphology, it does not demonstrate any preferential association between wall vesicles and microtubules thus suggesting that microtubules are only indirectly involved in wall vesicle transport. Freeze substitution is superior to conventional fixation for analysis of the Golgi apparatus because it uniquely reveals both differentiation of a specific single cisterna in each Golgi body and greater differences in membrane thicknesses throughout the endomembrane system.     

The cellular parameters of leaf development in tobacco: a clonal analysis

The cellular parameters of leaf development in tobacco (Nicotiana tabacum L.) have been characterized using clonal analysis, an approach that provides unequivocal evidence of cell lineage. Our results indicate that the tobacco leaf arises from a group of around 100 cells in the shoot apical meristem. Each of these cells contributes to a unique longitudinal section of the axis and transverse section of the lamina. This pattern of cell lincage indicates that primordial cells contribute more or less equally to the growth of the axis, in contrast to the more traditional view of leaf development in which the leaf is pictured as arising from a group of apical initials. Clones induced prior to the initiation of the lamina demonstrate that the subepidermal layer of the lamina arises from at least six files of cells. Submarginal cells usually divide with their spindles parallel to the margin, and therefore contribute relatively little to the transverse expansion of the lamina. During the expansion of the lamina the orientation and frequency of cell division are highly regulated, as is the duration of meristematic growth. Initially, cell division is polarized so as to produce lineages that are at an oblique angle to the midrib; later cell division is in alternating perpendicular planes. The distribution of clones generated by irradiation at various stages of development indicates that cell division ceases at the tip of the leaf when the leaf is about one tenth its final size, and then ceases in progressively more basal regions of the lamina. Variation in the mutation frequency within the lamina reflects variation in the frequency of mitosis. Prior to the mergence of the leaf the frequency of mutation is maximal near the tip of the leaf and extremely low at its base; after emergence, the frequency of mutation increases at the base of the leaf. In any given region of the lamina the frequency of mutation is highest in interveinal regions, and is relatively low near the margin. Thus, both the orientation and frequency of cell division at the leaf margin indicate that this region plays a minor role in the growth of the lamina.Abbreviation MF mutation frequency     

Cell size and chloroplast size in relation to chloroplast replication in light-grown wheat leaves

As part of an investigation into the control of chloroplast replication the number and size of chloroplasts in mesophyll cells was examined in relation to the size of the cells. In first leaves of Triticum aestivum L. and T. monococcum L. the number of chloroplasts in fully expanded mesophyll cells is positively correlated with the plan area of the cells. The linear relationship between chloroplast number per cell and cell plan area is also consistent over a fivefold range of cell size in isogenic diploid and tetraploid T. monococcum. In T. aestivum the chloroplast number per unit cell plan area varies among cells in relation to the size of the chloroplasts. Those cells containing chloroplasts with a relatively small face area have a correspondingly higher density of chloroplasts, and consequently, the total chloroplast area per unit cell plan area is very similar in all the cells. The results indicate that the proportion of the cell surface area covered by chloroplasts is precisely regulated, and that this is achieved during cell development by growth and replication of the chloroplasts.     

Control of the rate of cell enlargement: Excision,wall relaxation,and growth-induced water potentials

A new guillotine thermocouple psychrometer was used to make continuous measurements of water potential before and after the excision of elongating and mature regions of darkgrown soybean (Glycine max L. Merr.) stems. Transpiration could not occur, but growth took place during the measurement if the tissue was intact. Tests showed that the instrument measured the average water potential of the sampled tissue and responded rapidly to changes in water potential. By measuring tissue osmotic potential ( _s ), turgor pressure ( _p ) could be calculated. In the intact plant, _s and _p were essentially constant for the entire 22 h measurement, but _s was lower and _p higher in the elongating region than in the mature region. This caused the water potential in the elongating region to be lower than in the mature region. The mature tissue equilibrated with the water potential of the xylem. Therefore, the difference in water potential between mature and elongating tissue represented a difference between the xylem and the elongating region, reflecting a water potential gradient from the xylem to the epidermis that was involved in supplying water for elongation. When mature tissue was excised with the guillotine, _s and _p did not change. However, when elongating tissue was excised, water was absorbed from the xylem, whose water potential decreased. This collapsed the gradient and prevented further water uptake. Tissue _p then decreased rapidly (5 min) by about 0.1 MPa in the elongating tissue. The _p decreased because the cell walls relaxed as extension, caused by _p , continued briefly without water uptake. The _p decreased until the minimum for wall extension (Y) was reached, whereupon elongation ceased. This was followed by a slow further decrease in Y but no additional elongation. In elongating tissue excised with mature tissue attached, there was almost no effect on water potential or _p for several hours. Nevertheless, growth was reduced immediately and continued at a decreasing rate. In this case, the mature tissue supplied water to the elongating tissue and the cell walls did not relax. Based on these measurements, a theory is presented for simultaneously evaluating the effects of water supply and water demand associated with growth. Because wall relaxation measured with the psychrometer provided a new method for determining Y and wall extensibility, all the factors required by the theory could be evaluated for the first time in a single sample. The analysis showed that water uptake and wall extension co-limited elongation in soybean stems under our conditions. This co-limitation explains why elongation responded immediately to a decrease in the water potential of the xylem and why excision with attached mature tissue caused an immediate decrease in growth rate without an immediate change in _p Abbreviations and symbols L tissue conductance for water - m wall extensibility - Y average yield threshold (MPa) - _o water potential of the xylem - _p turgor pressure - _s osmotic potential - _w water potential of the elon gating tissue     

Immunofluorescent and calcofluor white staining of developing tracheary elements inZinnia elegans L. Suspension cultures

Summary Xylogenesis has been studied in primary suspension cultures ofZinnia elegans L.: The wall patterns produced in culture closely resemble those described for intact tissues (annular, spiral, reticulate, scalariform, pitted). Using fluorescence microscopy and immuno-cytochemical techniques we have followed both the changes in wall deposition and microtubule organization during xylogenesis. Calcofluor white has been used to detect secondary wall deposition before it can be observed using either phase contrast or polarization optics. The development of tracheary elements can be divided into three stages: 1. microtubules grouped into bands without secondary wall deposition evident; 2. groups of microtubules subtending wall material only visible using Calcofluor white; 3. a complex microtubule pattern reflected by well developed wall thickenings detected using Calcofluor, phase contrast and polarization optics.     

The flagellar apparatus of the golden algaSynura uvella: Four absolute orientations

Summary Flagellated vegetative cells of the colonial golden algaSynura uvella Ehr, were examined using serial sections. The two flagella are nearly parallel as they emerge from a flagellar pit near the apex of the cell. The photoreceptor is restricted to swellings on the flagella in the region where they pass through the apical pore in the scale case and the swellings are not associated with the cell membrane or an eyespot. A unique ring-like structure surrounds the axonemes of both flagella at a level just above the transitional helix. The basal bodies are interconnected by three striated, fibrous bands. Four short (<100 nm) microtubules lie between the basal bodies at their proximal ends. Two rhizoplasts extend down from the basal bodies and separate into numerous fine striated bands which lie over the nucleus. Three- and four-membered microtubular roots arise from the rhizoplasts and extend apically together. As the roots reach the cell anterior, the three-membered root bends and curves clockwise to form a large loop around the flagella; the four-membered root bends anticlockwise and terminates under the distal end of the three-membered root as it completes the loop. There are four absolute orientations, termed Types 1–4, in which the flagellar apparatus can occur. With each orientation type the positions of the Golgi body, nucleus, rhizoplasts, chloroplasts and microtubular roots change with respect to the flagella, basal bodies and photoreceptor. Two new basal bodies appear in pre-division cells, and three short microtubules appear in a dense substance adjacent to each new basal body. Based upon the positions of new pre-division basal bodies, a hypothesis is proposed to explain why there are four orientations and how they are maintained through successive cell divisions.     

Large scale selection of aluminum-resistant mutants from plant cell culture: expression and inheritance in seedlings

Summary A large number of aluminum-resistant variants, selected from non-mutagenized homozygous diploid cell cultures of Nicotiana plumbaginifolia Viv., are characterized. Of 115 variants cloned and reselected from single cells, 67 retained Al resistance in callus cultures after 6–9 months of growth in the absence of Al. There was no association between Al resistance and callus growth in the absence of Al, suggesting that the Al-resistant phenotype is not detrimental in the absence of Al challenge and that Al resistance is not the result of increased vigor. Plants regenerated from initially resistant callus lines that subsequently lost their resistance failed, with one exception, to transmit resistance to their seedling progeny. Fertile plants were regenerated from 40 of the 67 variants that retained stable Al resistance in callus culture. All 40 transmitted Al resistance to their seedling progeny (selfed and backcrossed) in segregation ratios expected for a single dominant mutation. The selfed progeny of many variants also segregated for recessive lethal mutations which were attributed to independent mutations that occurred during cell culture.