The Azolla, Anabaena azollae Relationship: II. Localization of Nitrogenase Activity as Assayed by Acetylene Reduction

Anaerobic (microaerophilic) acetylene reduction by Azolla caroliniana Willd. was dependent on light and saturated at approximately 450 foot candles. Maximum rates of acetylene reduction were 60 nmoles/mg chlorophyll minute. However, rates of 25 to 30 nmoles/mg chlorophyll minute were more common.     

The Azolla-Anabaena azollae Relationship : XI. PHYCOBILIPROTEINS IN THE ACTION SPECTRUM FOR NITROGENASE-CATALYZED ACETYLENE REDUCTION

Visible absorption spectra are presented for the Azolla caroliniana Willd.-Anabaena azollae Strass. association and the individual partners. Although absorption by the phycobiliproteins of the endophytic cyanobacterium clearly complements the absorption by the fern pigments, their contribution to the absorption spectrum of the association is effectively concealed by the preponderance of the Azolla pigments. Action spectra for nitrogenase-catalyzed C₂H₂ reduction in both the Azolla-Anabaena association and the endophytic Anabaena demonstrate that quanta absorbed by the phycobiliproteins is as effective as that absorbed by chlorophyll a in driving this photosystem I-linked process. Under anaerobic conditions, the inhibition of photosystem II activity by 3-(3,4-dichlorophenyl)-1,1-dimethylurea, diuron did not selectively decrease the relative quantum yields in the region of phycobiliprotein absorption. At the well-below saturating light intensities used for the action spectra studies, the absolute rates of C₂H₂ reduction were increased uniformly via respiratory-linked processes under aerobic conditions. The occurrence of phycobiliproteins in heterocysts of the endophytic Anabaena was demonstrated using fluorescence microscopy of intact filaments. Fluorescence micrographs of Anabaena cylindrica filaments are presented for comparison.     

Azolla-Anabaena azollae Relationship: V. N(2) Fixation, Acetylene Reduction, and H(2) Production

In order to characterize the reactions catalyzed by nitrogenase in the Azolla-Anabaena association, ¹⁵N₂ fixation, C₂H₂ reduction, and ATP-dependent H₂ production were measured in both the Azolla-Anabaena complex and in the alga isolated from the complex.     

Azolla-Anabaena Relationship: VIII. Photosynthetic Characterization of the Association and Individual Partners

Photosynthesis in the Azolla-Anabaena association was characterized with respect to photorespiration, early products of photosynthesis, and action spectra. Photorespiration as evidenced by an O₂ inhibition of photosynthesis and an O₂-dependent CO₂ compensation concentration was found to occur in the association, and endophyte-free fronds, but not in the endophytic Anabaena. Analysis of the early products of photosynthesis indicated that both the fern and cyanobacterium fix CO₂ via the Calvin cycle. The isolated endophytic Anabaena did not release significant amounts of amino acids synthesized from recently fixed carbon. The action spectra for photosynthesis in the Azolla-Anabaena association indicated that the maximum quantum yield is between 650 and 670 nanometers, while in the endophyte the maximum is between 580 and 640 nanometers. Although the endophytic cyanobacterium is photosynthetically competent, any contribution it makes to photosynthesis in the intact association was not apparent in the action spectrum.     

The Azolla, Anabaena azollae Relationship: I. Initial Characterization of the Association

Cultures of Azolla caroliniana Willd. free of the symbiotic blue-green alga, Anabaena azollae, were obtained by treatment of Azolla fronds with a regimen of antibiotics. These symbiontfree plants can be maintained only on medium containing a combined nitrogen source.     

Biosynthesis of Sucrose and Mannitol as a Function of Leaf Age in Celery (Apium graveolens L.)

In celery (Apium graveolens L.), the two major translocated carbohydrates are sucrose and the acyclic polyol mannitol. Their metabolism, however, is different and their specific functions are uncertain. To compare their roles in carbon partitioning and sink-source transitions, developmental changes in ¹⁴CO₂ labeling, pool sizes, and key enzyme activities in leaf tissues were examined. The proportion of label in mannitol increased dramatically with leaf maturation whereas that in sucrose remained fairly constant. Mannitol content, however, was high in all leaves and sucrose content increased as leaves developed. Activities of mannose-6-P reductase, cytoplasmic and chloroplastic fructose-1,6-bisphosphatases, sucrose phosphate synthase, and sucrose synthase increased with leaf maturation and decreased as leaves senesced. Ribulose bisphosphate carboxylase and nonreversible glyceraldehyde-3-P dehydrogenase activities rose as leaves developed but did not decrease. Thus, sucrose is produced in all photosynthetically active leaves whereas mannitol is synthesized primarily in mature leaves and stored in all leaves. Onset of sucrose export in celery may result from sucrose accumulation in expanding leaves, but mannitol export is clearly unrelated to mannitol concentration. Mannitol export, however, appears to coincide with increased mannitol biosynthesis. Although mannitol and sucrose arise from a common precursor in celery, subsequent metabolism and transport must be regulated separately.     

Calcium Status of Yellow Lupin Symbiosomes as a Potential Regulator of Their Nitrogenase Activity: The Role of the Peribacteroid Membrane

The capacity of symbiosomes from yellow lupin root nodules for active Ca²⁺uptake and the sensitivity of their nitrogenase activity to a disturbance of the symbiotic Ca partition were investigated. The experiments carried out on the isolated symbiosomes and the peribacteroid membrane (PBM) vesicles, using Ca²⁺indicators arsenazo III and chlorotetracycline, and the cytochemical Ca visualization with potassium pyroantimonate (PA) provided evidence that an Mg-ATP-energized pump, most likely Mg²⁺-dependent Ca²⁺-ATPase catalyzing the active transport of Ca²⁺from the cytosol of the plant cell into the symbiosomes across the PBM, functions on this membrane. Depleting the symbiosomes of Ca both in vivoandin vitroby treating the intact nodules of yellow lupin root or the purified symbiosomes isolated from the latter with EGTA and Ca²⁺-ionophore A23187 substantially decreased their nitrogenase activity. The inhibitory effect of calcium deficit in the symbiosomes was not reversed by the addition of calcium to the incubation medium containing the plant tissues under study and was even enhanced under these conditions. The nitrogenase activity of the isolated symbiosomes not experiencing calcium deficit was also inhibited by the addition of relatively high concentrations of exogenous calcium to the incubation medium. These results seem to give evidence that the calcium status of nodule symbiosomes from yellow lupin roots controls their nitrogenase activity. The data obtained suggest that both Ca²⁺transport on PBM and the low passive permeability of this membrane for the given cation play the key role in such a control.     

四棱豆(Psophocarpus tetragonolobus)酰脲相对丰度与固氮酶活力的关系

四棱豆根瘤固氮酶活力日变化呈双峰曲线,两个峰分别出现在14:30和20:00;其固氮酶活力日变化与叶片酰脲含量变化的相关系数为0.67,且与根、茎、叶的酰脲相对丰度(URA)相关系数分别为0.59,0.61,0.76。在个体发育过程中,根瘤固氮酶活力与叶片酰脲含量以及酰脲相对丰度之间呈极显著相关。贮存在30℃条件下5 h不影响植物材料的酰脲相对丰度。四棱豆叶片酰脲相对丰度可作为估价根瘤固氮酶活力的一个指标。     

Hydrolytic Enzymes of Euglena gracilis: Characterization and Activity as a Function of Culture Age and Carbon Deprivation*†

SYNOPSIS. Optimal assay conditions are described for 8 hydrolases of Euglena gracilis var. bacillaris, SM-L1 (streptomycinbleached) strain, 7 of which have an acid pH-optimum. Acid phosphatase, β-galactosidase, β-glucosidase, β-fucosidase, cathepsin D, RNase, DNase, and an esterase are active in cell homogenates. Amylase has very low activity, and β-glucuronidase, arylsulfatase, β, N-acetyl-glucosaminidase, α-fucosidase, and α- and β-mannosidase are inactive. Hydrolase activity increases as a culture proceeds from the midexponential to the late stationary-phase of growth, being most pronounced in the case of β-glucosidase. In cultures deprived of a utilizable carbon source, the specific activities of the hydrolases (per mg total protein or dry weight) increase. When expressed on a per cell basis, however, the activities of DNase decrease while those of β-galactosidase, cathepsin D, and RNase increase. The hydrolases appear to be involved in the adaptation of Euglena to the metabolic demands imposed by different conditions of growth.     

Elevation-Related Variation in Leaf Stomatal Traits as a Function of Plant Functional Type: Evidence from Changbai Mountain,China

Understanding the variation in stomatal characteristics in relation to climatic gradients can reveal the adaptation strategies of plants, and help us to predict their responses to future climate changes. In this study, we investigated stomatal density (SD) and stomatal length (SL) in 150 plant species along an elevation gradient (540–2357 m) in Changbai Mountain, China, and explored the patterns and drivers of stomatal characteristics across species and plant functional types (PFTs: trees, shrubs, and herbs). The average values of SD and SL for all species combined were 156 mm^–2 and 35 µm, respectively. SD was higher in trees (224 mm^–2) than in shrubs (156 mm^–2) or herbs (124 mm^–2), and SL was largest in herbs (37 µm). SD was negatively correlated with SL in all species and PFTs (P<0.01). The relationship between stomatal characteristics and elevation differed among PFTs. In trees, SD decreased and SL increased with elevation; in shrubs and herbs, SD initially increased and then decreased. Elevation-related differences in SL were not significant. PFT explained 7.20–17.6% of the total variation in SD and SL; the contributions of CO₂ partial pressure (), precipitation, and soil water content (SWC) were weak (0.02–2.28%). Our findings suggest that elevation-related patterns of stomatal characteristics in leaves are primarily a function of PFT, and highlight the importance of differences among PFTs in modeling gas exchange in terrestrial ecosystems under global climate change.     

The Relationship Between the Distribution of Periclinal Cell Divisions in the Shoot Apex and Leaf Initiation

Periclinal cell divisions in vegetative shoot apices of Pisumand Silene were recorded from serial thin sections by mappingall the periclinal cell walls formed less than one cell cyclepreviously. The distribution of periclinal divisions in theapical domes corresponded to the distributions subsequentlyoccurring in the apices when the young leaf primordia were forming.In Pisum, periclinal divisions were almost entirely absent fromthe I₁ region of the apical dome for half a plastochron justafter the formation of a leaf primordium and appeared, simultaneouslyover the whole of the next potential leaf site, about half aplastochron before the primordium formed. In Silene periclinaldivisions seemed to always present in the apical dome at thepotential leaf sites and also round the sides of the dome wherethe ensheathing leaf bases were to form. Periclinal divisionstherefore anticipated the formation of leaf primordia by occuring,in Pisum about one cell cycle and in Silene two or more cellcycles, before the change in the direction of growth or deformationof the surface associated with primordial initiation. Pisum, Silene, planes of cell division, orientation of cell walls, leaf primordia, shoot apical meristem, plastochron     

Calcium in the Control of Nitrogenase Activity in Vicia faba L. Root Nodules: Calcium Release from Symbiosomes and the Accompanying Decrease in Their Nitrogenase Activity Is Blocked by Verapamil

Treatment of root nodules or symbiosomes isolated from them with calcium chelator EGTA alone or together with calcium ionophore A23187 for 3 h under microaerophilic conditions considerably decreased their nitrogenase activity (NA). Under these experimental conditions, cytochemical electron-microscopic analysis revealed considerable calcium depletion of symbiosomes in the infected nodule cells treated with EGTA and A23187. Ca²⁺ channel blockers, verapamil and ruthenium red, inhibited EGTA-induced Ca²⁺ release from symbiosomes. In this case, NA insignificantly increased in the whole nodules and reached its initial level in symbiosomes. The experiments on isolated symbiosomes with arsenazo III, a Ca²⁺ indicator, demonstrated that verapamil inhibited Ca²⁺ release from them induced by valinomycin in the presence of K⁺ ions. These data suggest the presence on the peribacteroid membrane of a verapamil-sensitive transporter responsible for Ca²⁺ release from symbiosomes. A possible role of this transporter in the interaction between symbiotic partners in the infected cells of root nodules is discussed.     

吡啶锰配合物诱导肿瘤细胞死亡的作用及对线粒体功能的影响

研究新合成的小分子吡啶锰配合物Adpa-Mn（III）（[（Adpa）Mn（μ2-O）2Mn（Adpa）]PF6.8H2O（Adpa=bis（2-pyridylmethyl）amino-2-propionic acid））的抗肿瘤作用,初步探索其抗肿瘤的机制。MTT分析Adpa-Mn（III）对细胞活性的影响;活细胞工作站观察GFP荧光标记组蛋白HeLa细胞的细胞核形态,MDC染色以及GFP-LC3质粒转染,探讨细胞死亡的方式;JC-1染色检测线粒体膜电位;Fluo-3-AM和DCFH-DA荧光探针分别检测细胞中Ca^2＋和ROS的含量。结果发现,Adpa-Mn（III）剂量依赖性地抑制细胞活性;给药后细胞核出现固缩、片段化;自噬小泡增多,GFP-LC3荧光强度增强;线粒体膜电位下降;细胞内Ca^2＋发生超载,ROS含量升高。由此,Adpa-Mn（III）可抑制肿瘤细胞活性,其机制与引起线粒体膜电位下降、增加ROS生成及诱导细胞的死亡有关,同时胞内Ca^2＋超载也参与了该作用。这些数据显示,Adpa-Mn（III）具有成为抗肿瘤先导金属配合物的潜在可能性。     

The Influence of Light and Oxygen on Nitrogenase Activity in the Lichen Stereocaulon paschale

Thalli of the lichen Slereocaulon paschale (L.) Fr. were prctreated in the light (light activated) or in the dark (dark starved). In short-time experiments with both light activated arid dark starved thalli, the nitrogenasc activity was higher in the light than in the dark, Light activated thalli had a very much higher rale of C₂H₂ reduction than dark starved thalli, both in the light and in the dark. The dark starved lhalli showed increasing nilrogenase activity when incubated in the light. Either light or oxygen was necessary for nitrogenase activity in light activated thalli. and up to about 10kPa oxygen they showed additive effects. Both in the light and in the dark the nitrogenase activity decreased when the oxygen partial pressure was lower than in normal air. The experimental data thus showed a short-term effect of light on nilrogenase activity by provision of ATP and reductant, and a long term effect probably by build up of reserves that were later utilized. Any immediate effect of photorespiration on nitrogenase activity could not be found in light activated thalli.     

Comparison between a Stable NaCl-Selected Nicotiana Cell Line and the Wild Type : K, Na, and Proline Pools as a Function of Salinity

An NaCl-resistant line has been developed from suspension-cultured tobacco cells (Nicotiana tabacum/gossii) by stepwise increases in the NaCl concentration in the medium. Resistance showed stability through at least 24 generations in the absence of added NaCl.

Above an external NaCl concentration of 35 millimolar, proline concentration in the selected cells rose steeply with external NaCl, particularly so above 100 millimolar NaCl. Proline accumulation in the wild type was far slighter. Selected cells which had been grown for 24 generations in the absence of added NaCl accumulated proline strongly on re-exposure to NaCl medium, indicating stability of this character. Proline accumulation was fully reversible with a half-time of about 6 hours. When selected cells were transferred sequentially to lower and lower NaCl concentrations, their proline content fell to the level corresponding to the new NaCl concentration. The NaCl-selected cells responded to water stress (i.e. added mannitol) by accumulating markedly more proline than did the wild type.

The addition of Ca²⁺ to the growing and rinsing media minimized Na⁺ and K⁺ binding in the Donnan free space of cell walls and thus allowed assessment of intracellular Na⁺ and K⁺. In both cell types, internal Na⁺ content rose steadily as a function of external NaCl concentration. In the course of 7 days in NaCl media, the wild type cells lost a considerable part of their K⁺ content, the extent of the loss increasing with rise in external NaCl concentration. The selected cells, by contrast, lost no K⁺ at external NaCl concentrations below 50 millimolar external NaCl, and at higher concentrations lost less than the wild type.

     

Studies on Chromophore Coupling in Isolated Phycobiliproteins: II. Picosecond Energy Transfer Kinetics and Time-Resolved Fluorescence Spectra of C-Phycocyanin from Synechococcus 6301 as a Function of the Aggregation State

The fluorescence kinetics of C-Phycocyanin in the monomeric, trimeric, and hexameric aggregation states has been measured as a function of the emission wavelength with picosecond resolution using the single-photon timing technique. All the decay curves measured at the various emission wavelengths were analyzed simultaneously by a global data analysis procedure. A sum of four exponentials was required to fit the data for the monomers and trimers. Only in the case of the hexamers, a three-exponential model function proved to be nearly sufficient to describe the experimental decays. The lifetime of those fluorescence components reflecting energy transfer decreased with increasing aggregation. This is due to the increased number of efficient acceptor molecules next to a donor in the higher aggregates. In all aggregates the shortest-lived component, ranging from 50 ps for monomer to 10 ps for hexamers, is observed as a decay term (positive amplitude) at short emission wavelength. At long emission wavelength it turns into a rise term (negative amplitude). The lifetime of a second ps-component ranges from 200 ps for monomers to 50 ps for hexamers. The long-lived (ns) fluorescence is inhomogeneous in monomers and trimers, showing two lifetimes of ~0.6 and 1.3 ns. The latter one carries the larger amplitude. The amplitudes of the kinetic components in the fluorescence decays are presented as time-resolved component spectra. A theoretical model has been derived to rationalize the observed fluorescence kinetics. Using symmetry arguments, it is shown that the fluorescence kinetics of C-Phycocyanin is expected to be characterized by three exponential kinetic components, independent of the aggregation state. An analytical expression is derived, which allows us to gain a detailed understanding of the origin of the different kinetic components and their associated time-resolved spectra. Numerical calculations of time-resolved spectra are compared with the experimental data.     

The Development and Use of a Flow-through Apparatus for Measuring Nitrogenase Activity and Photosynthesis in Field Crops

A flow-through apparatus for measuring nitrogenase activityin field-grown crops is described. The apparatus was used tomeasure nitrogenase activity in a sainfoin crop throughout agrowth season. A marked seasonal variation was observed whichwas partly the consequence of defoliation. The acetylene-basedestimate of the nitrogen fixed annually by sainfoin was less(146 kg N ha^–1) than the nitrogen accumulated in the shoots.Reasons for this difference are discussed with special emphasison the validity of the technique and alternative sources ofnitrogen. There were no treatment differences between the ratesof growth of either sainfoin or lucerne when grown with or withoutfertilizer nitrogen in the field. This result is in marked contrastwith results from a glasshouse experiment where sainfoin plantsutilizing nitrate-nitrogen exhibited faster growth rates thansimilar plants that were solely dependent on symbiotic nitrogenfixation. Key words: Nitrogenase, photosynthesis, field-technique, sainfoin, lucerne     

Grass Leaf Elongation Rate as a Function of Developmental Stage and Temperature: Morphological Analysis and Modelling

Elongation of successive leaves was measured following defoliationof tall fescue plants in controlled environments. Measurementswere made under constant temperatures of 24 °C and 14 °C,and after temperature changes from 24 to 14 °C andvice versa.A morphological analysis of the growing leaf was made from thetime it was 1 mm long until it was fully elongated. The timeelapsed from initiation until the leaf was 1 mm long was estimated.Young leaves less than 1.5 mm long elongated slowly at a constantleaf elongation rate (LER). By extrapolating this LER back toleaf initiation from the apex it was calculated that elongationlasted 42.5 d at 24 °C and 51 d at 14 °C. Lengths ofthe division zone (DZ) and the extension-only zone (E-OZ) increasedto a maximum and then decreased during leaf development. Temperaturechange had an immediate effect on LER but the response varieddepending on the direction of the temperature change. To describethese different features, an empirical model of DZ and E-OZwas designed. Its five parameters were optimized at constanttemperature. The model was then used to simulate the LER ofplants subjected to temperature changes. Instant and lastingeffects of the initial temperature on mean LER in plants transferredfrom 14 to 24 °C andvice versawere well simulated. It wasconcluded that the major reason for differences was due to thegrowth stage (DZ and E-OZ lengths) at which the changes occurredat both temperatures.Copyright 1999 Annals of Botany Company Festuca arundinaceaSchreb., tall fescue, growth zone, division zone.     

Review: The Dynamics of the Nuclear Lamins during the Cell Cycle— Relationship between Structure and Function

The nuclear lamins are members of the intermediate filament (IF) family of proteins. The lamins have an essential role in maintaining nuclear integrity, as do the other IF family members in the cytoplasm. Also like cytoplasmic IFs, the organization of lamins is dynamic. The lamins are found not only at the nuclear periphery but also in the interior of the nucleus, as distinct nucleoplasmic foci and possibly as a network throughout the nucleus. Nuclear processes such as DNA replication may be organized around these structures. In this review, we discuss changes in the structure and organization of the nuclear lamins during the cell cycle and during cell differentiation. These changes are correlated with changes in nuclear structure and function. For example, the interactions of lamins with chromatin and nuclear envelope components occur very early during nuclear assembly following mitosis. During S-phase, the lamins colocalize with markers of DNA replication, and proper lamin organization must be maintained for replication to proceed. When cells differentiate, the expression pattern of lamin isotypes changes. In addition, changes in lamin organization and expression patterns accompany the nuclear alterations observed in transformed cells. These lamin structures may modulate nuclear function in each of these processes.