Can genotypes of soybean (Glycine max) selected for nitrate tolerance provide good "models" for studying the mechanism of nitrate inhibition of nitrogenase activity?

In soybeans ( Glycine max L. Merr.), high levels of soil nitrate inhibit N₂ fixation, and nitrate-tolerant symbioses have been identified within a chemically mutagenized line of cv. Bragg denoted nts382 and within the line K466, a genotype representative of a number of Korean soybean cultivars. The genotypes nts382 and K466 were examined to see if they could be used as a model system for studying the mechanism responsible for the short-term (i.e. 3-day) inhibition of specific nitrogenase activity, especially the mechanism behind the greater O₂ limitation of nodule metabolism that is characteristic of nitrate inhibition of N₂ fixation in soybean. In nts382, total nitrogenase activity (TNA = H₂ production in Ar:O₂) was inhibited to a lesser degree (48% of control) relative to Bragg (30% of control), and the nitrate-treated symbioses showed less of an O₂ limitation of nodule metabolism in nts382 than in Bragg. However, the relative proportion of O₂ limitation to the total nitrate inhibition was similar (40 and 41%) in nts382 and Bragg, respectively. Therefore, the nts382 symbioses may be useful in elucidating the general mechanism for down-regulation of nitrogenase activity in soybean, but would not be a useful model system for studying the control of O₂-limited metabolism following nitrate exposure. The effects of nitrate on TNA and on the degree of O₂ limitation of nodule metabolism were the same in K466 and a reference cultivar Maple Arrow. Consequently, the tolerance of K466 to nitrate reported previously was attributed to the ability of this symbiosis to maintain nodule biomass in the presence of nitrate, not to any ability to maintain specific nitrogenase activity in the presence of nitrate.     

What triggers the regulation of nitrogenase activity in forage legume nodules after defoliation?

The dramatic decrease in nitrogenase activity after the defoliation of forage legumes has been recognized for a long time; however, the underlying mechanisms are not understood yet. The impact of current photosynthesis can be excluded. The precise role of carbohydrate availability is still unclear and remains to be established. From current knowledge we can conclude that, after defoliation, nitrogenase activity in legume nodules is down-regulated by a variable oxygen diffusion resistance. The triggering elements are not known; there is, however, increasing evidence that the plant's demand for symbiotically fixed nitrogen plays an important role. The possibility is here discussed that, after defoliation, a nitrogen feedback mechanism regulates nitrogenase activity through a variable oxygen diffusion resistance in the nodules.     

Can genotypes of soybean (Glycine max) selected for nitrate tolerance provide good “models” for studying the mechanism of nitrate inhibition of nitrogenase activity?

In soybeans ( Glycine max L. Merr.), high levels of soil nitrate inhibit N₂ fixation, and nitrate-tolerant symbioses have been identified within a chemically mutagenized line of cv. Bragg denoted nts382 and within the line K466, a genotype representative of a number of Korean soybean cultivars. The genotypes nts382 and K466 were examined to see if they could be used as a model system for studying the mechanism responsible for the short-term (i.e. 3-day) inhibition of specific nitrogenase activity, especially the mechanism behind the greater O₂ limitation of nodule metabolism that is characteristic of nitrate inhibition of N₂ fixation in soybean. In nts382, total nitrogenase activity (TNA = H₂ production in Ar:O₂) was inhibited to a lesser degree (48% of control) relative to Bragg (30% of control), and the nitrate-treated symbioses showed less of an O₂ limitation of nodule metabolism in nts382 than in Bragg. However, the relative proportion of O₂ limitation to the total nitrate inhibition was similar (40 and 41%) in nts382 and Bragg, respectively. Therefore, the nts382 symbioses may be useful in elucidating the general mechanism for down-regulation of nitrogenase activity in soybean, but would not be a useful model system for studying the control of O₂-limited metabolism following nitrate exposure. The effects of nitrate on TNA and on the degree of O₂ limitation of nodule metabolism were the same in K466 and a reference cultivar Maple Arrow. Consequently, the tolerance of K466 to nitrate reported previously was attributed to the ability of this symbiosis to maintain nodule biomass in the presence of nitrate, not to any ability to maintain specific nitrogenase activity in the presence of nitrate.     

Does oxygen limit nitrogenase activity in soybean exposed to elevated CO2?

Soybean (Glycine max L. Merr) plants grown under control (360 µmol mol^?1) or elevated CO₂ concentration (800 µmol mol^?1) from 33 to 42 d after sowing were assayed for various components of in vivo nitrogenase activity to test the hypothesis that increasing carbohydrate supply to nodules would increase the potential (i.e. O₂ saturated) nitrogenase activity and impose a more severe O₂ limitation on both nodule metabolism and total nitrogenase activity. Within 51 h of elevated CO₂ treatment, significant increases relative to control plants were seen in total nitrogenase activity expressed per plant. After 6 d of elevated CO₂, the total nitrogenase activity per plant was 18% higher than that in control. This was attributed to an initial increase in nodule size, and a subsequent increase in nodule number following plant exposure to elevated CO₂. However, after 9 d of elevated CO₂, the potential and total nitrogenase activities per gram nodule dry weight were lower, not higher than corresponding values in plants in the control treatment. These results did not support the hypothesis. It was concluded that the metabolic capacity of the control nodules were not limited by carbohydrate supply, at least at the assay temperatures employed here.     

Influence of nitrate on uptake of ammonium by nitrogen-depleted soybean: is the effect located in roots or shoots?

In non-nodulated soybean [Glycine max (L.) Merrill cv. Ransom]plants that were subjected to 15 d of nitrogen deprivation inflowing hydroponic culture, concentrations of nitrogen declinedto 1.0 and 1.4mmol Ng^–1 dry weight in shoots and roots,respectively, and the concentration of soluble amino acids (determinedas primary amines) declined to 40µmol g^–1 dry weightin both shoots and roots. In one experiment, nitrogen was resuppliedfor 10 d to one set of nitrogen-depleted plants as 1.0 mol m^–3NH₄⁺ to the whole root system, to a second set as 0.5 mol m^–3NH₄⁺ plus 0.5 mol m^–3 NO₃^– to the whole root system,and to a third set as 1.0 mol m^–3 NH₄⁺ to one-half ofa split-root system and 1.0 mol m^–3 NO₃^– to theother half. In a second experiment, 1.0 mol m^–3 of nitrogenwas resupplied for 4 d to whole root systems in NH₄⁺ : NO₃^–ratios of 1:0, 9:1, and 1:1. Nutrient solutions were maintainedat pH 6.0. When NH₄⁺ was resupplied in combination with NO₃^– to thewhole root system in Experiment I, cumulative uptake of NH₄⁺for the 10 d of resupply was about twice as great as when NH₄⁺was resupplied alone. Also, about twice as much NH₄⁺ as NO₃^–was taken up when both ions were resupplied to the whole rootsystem. When NH₄⁺ and NO₃^– were resupplied to separatehalves of a split-root system, however, cumulative uptake ofNH₄⁺ was about half that of NO₃^–. The uptake of NH₄⁺,which is inhibited in nitrogen-depleted plants, thus is facilitatedby the presence of exogenous NO₃^–, and the stimulatingeffect of NO₃^– on uptake of NH₄⁺ appears to be confinedto processes within root tissues. In Experiment II, resupplyof nitrogen as both NH₄⁺ and NO₃^– in a ratio of either1:1 or 9:1 enhanced the uptake of NH₄⁺. The enhancement of NH₄⁺uptake was 1.8-fold greater when the NH₄⁺: NO₃^–-resupplyratio was 1:1 than when it was 9:1; however, only 1.3 timesas much NO₃^– was taken up by plants resupplied with the1 :1 exogenous ratio. The effect of NO₃^– on enhancementof uptake of NH₄⁺ apparently involves more than net uptake ofNO₃^– itself and perhaps entails an effect of NO₃^–uptake on maintenance of K⁺ availability within the plant. Theconcentration of K⁺ in plants declined slightly during nitrogendeprivation and continued to decline following resupply of nitrogen.The greatest decline in K⁺ concentration occurred when nitrogenwas resupplied as NH₄⁺ alone. It is proposed that decreasedavailability of K⁺ within the NH₄⁺-resup-plied plants inhibitedNH₄⁺ uptake through restricted transfer of amino acids fromthe root symplasm into the xylem. Key words: Ammonium, Glycine max, nitrate, nitrogen-nutrition, nitrogen stress, split-root cultures     

Carbon cost of nitrogenase activity in Frankia–Alnus incana root nodules

The carbon cost of nitrogenase activity was investigated to determine symbiotic efficiency of the actinorhizal root nodule symbiosis between the woody perennial Alnus incana and the soil bacterium Frankia. Respiration (CO₂ production) and nitrogenase activity (H₂ production) by intact nodulated root systems were continuously recorded in short-term assays in an open-flow gas exchange system. The assays were conducted in N₂:O₂, thus under N₂-fixing conditions, in all experiments except for one. This avoided the declines in nitrogenase activity and respiration due to N₂ deprivation that occur in acetylene reduction assays and during extended Ar:O₂ exposures in H₂ assays. Two approaches were used: (i) direct estimation of root and nodule respiration by removing nodules, and (ii) decreasing the partial pressure of O₂ from 21 to 15% to use the strong relationship between respiration and nitrogenase activity to calculate CO₂/H₂. The electron allocation of nitrogenase was determined to be 0.6 and used to convert the results into moles of CO₂ produced per 2e^– transferred by nitrogenase to reduction of N₂. The results ranged from 2.6 to 3.4mol CO₂ produced per 2e^–. Carbon cost expressed as gC produced per gN reduced ranged from 4.5 to 5.8. The result for this actinorhizal tree symbiosis is in the low range of estimates for N₂-fixing actinorhizal symbioses and crop legumes. Methodology and comparisons of root nodule physiology among actinorhizal and legume plants are discussed.     

The inhibitory effect of human interferon α on the generation of lymphokine-activated killer activity

Summary The generation of lymphokine-activated killer (LAK) activity and the proliferative response to human recombinant interleukin-2 (IL-2) were significantly reduced by the presence of human recombinant leukocyte interferon (IFN) in cultures of human peripheral blood mononuclear cells (PBMC). Mature natural killer (NK) cells can be depleted from PBMC with the toxic lysosomotropic agentl-leucine methyl ester. The generation of cytotoxic cells from lymphocytes depleted in leucine methyl ester was also inhibited by indicating that the IFN- effect is not limited to mature cytotoxic NK cells. Depletion of adherent cells from PBMC did not affect the suppression of LAK induction by IFN-. Surface marker analyses of Tac antigen and transferrin receptor (TfR) showed that the presence of IFN throughout the culture period significantly suppressed the typical increase in IL-2-induced Tac- and TfR-positive cells. In contrast, IFN treatment before and after IL-2 culture enhanced LAK cytotoxic activity. Therefore, combinations of these biological response modifiers for clinical use should take into account the dual effect of IFN on key features of the IL-2 response.This work was supported in part by USPHS grant CA34442. Y. T. is a UCLA visiting scientist from the Department of Surgery, School of Medicine, Tokai University, Kanagawa, Japan and recipient of a fellowship from the University of California Cancer Research Coordinating Committee. N. E. is a UCLA visiting scientist from the Department of Surgery, School of Medicine, Tohoku University, Sendai, Japan     

Can phosphorus limitation contribute to the maintenance of sex? A test of a key assumption

Why sex is so common remains unclear; what is certain is that the predominance of sex despite its profound costs means that it must confer major advantages. Here, we use elemental and nucleic acid assays to evaluate a key element of a novel, integrative hypothesis considering whether sex might be favoured because of differences in body composition between sexuals and asexuals. We found that asexual Potamopyrgus antipodarum, a New Zealand snail, have markedly higher bodily phosphorus and nucleic acid content per unit mass than sexual counterparts. These differences coincide with and are almost certainly linked to the higher ploidy of the asexuals. Our results are the first documented body composition differences between sexual and asexual organisms, and the first detected phenotypic difference between sexual and asexual P. antipodarum, an important natural model system for the study of the maintenance of sex. These findings also verify a central component of our hypothesis that competition between diploid sexuals and polyploid asexuals could be influenced by phosphorus availability.     

The effect of some ions on the activity of β-galactosidase and the inhibitory effect of tris (hydroxymethyl) aminomethane

The stimulating effect of phosphate and the inhibitory effect of tris-HCl on the activity of β-galactosidase inEscherichia coli was studied. The phosphate anion antagonizes the inhibitory effect of chloride. Since a similar effect is displayed by sulphate and arsenate no specific “stimulating” effect of phosphate can take place. The tris cation has also an inhibitory effect which is antagonized by univalent cations (K⁺). The resulting β-galactosidase activity reflects the antagonisms between cations and anions present in the reaction medium.     

Can charge recombination as caused by pH-dependent donor-side limitation in PS 2 account for high-energy-state quenching?

Schreiber and Neubauer (Photosynthesis Research 25: 279–293, 1990) have proposed a model which explains energy quenching by enhanced triplet formation as caused by charge recombination due to pH-dependent donor-side limitation. Quenching under these conditions is assumed to result from two mechanisms. Firstly, there is the withdrawal of excited states by charge recombination and formation of triplet states. Secondly, these triplet states can result in carotenoid triplets in the antenna which are supposed to quench excitons. Here, it is shown that quenching caused by both mechanisms can account for only about 25% of the experimentally observed energy quenching even under extremely favorable conditions. More likely, this number is less than 15%, as the contribution of the second step in the proposed triplet cycle is expected to be low as the life times of the carotenoid triplets are not long enough to cause the assumed quenching of excitons in the antenna.     

Can compartment models account for nonmonotonicity in pharmacokinetics?

The purpose of this paper is to show that for most of the compartment models used in pharmacokinetics, when initial conditions are zero except in one compartment, the time course of the corresponding state variable is monotonic decreasing. This statement is in contradiction with two related papers of Vaidhyanathan [1, 2], but is shown to be a consequence of theorems derived by Hearon [3].     

Effects of γ-irradiation on antioxidant activity in soybean seeds

There are some reports that low doses of γ-irradiation could induce antioxidant activities in plant material, including soybean. Irradiation, required for the inactivation of some pathogens and induction of mutations, may have adverse effects on sensorial, nutritional and antioxidant qualities. The effects of different γ-irradiation doses (100–200 Gy) on antioxidant properties of soybean seeds was investigated. In this study, we report the results obtained by analysis of antioxidant enzyme activities, reduced glutathione, malonyldialdehyde (MDA) and hydroxyl (HO⁻) radical quantities, soluble protein content, and total antioxidant activity in irradiated soybean seeds. Antioxidant enzyme activities were affected due to high irradiation intensity. Significant changes of total antioxidant activity and MDA and HO.quantities were observed only under the highest irradiation dose, with a 15.7% reduction in total antioxidant activity, MDA quantity increase of 21.6%, and HO⁻ radical quantity increase of 79.3% compared to the non-irradiated control. The total soluble protein content increased slightly.     

Can paternal preconceptional radiation account for the increase of leukaemia and non-Hodgkin's lymphoma in Seascale?

OBJECTIVE--To determine if the excess of leukaemia and non-Hodgkin''s lymphoma in Seascale is restricted to those born in the parish and whether it might be explained by the postulated relation with paternal preconceptional radiation. DESIGN--Comparison, separately for those born in the parish and those born elsewhere, of the numbers of these malignancies observed in Seascale with those expected on the basis of reference rates for England and Wales. Details of paternal radiation levels were sought for each case. SETTING--The parish of Seascale in west Cumbria. SUBJECTS--Residents of Seascale below age 25 years in the years 1951-91. MAIN OUTCOME MEASURES--The observed and expected numbers of cases of leukaemia and non-Hodgkin''s lymphoma within Seascale among those born there and among those born elsewhere. Also, the levels of any paternal preconceptional radiation associated with each case. RESULTS--A significant excess of leukaemia and non-Hodgkin''s lymphoma at ages 0-24 was found in Seascale in those who were born there (ratio of observed to expected cases 8.6 and 20.2 respectively; p < 0.01). This also applied to those not born there (7.2 and 16.5; p < 0.01), a group often regarded as not showing an excess. The estimates were then conservatively recalculated so as to overestimate the risks among those born in Seascale and underestimate them among those born elsewhere. On this basis the six cases in those born in Seascale compare with 0.38 expected (15.8; p < 0.001), of which two were associated with paternal preconceptional life-time levels of 100 mSv or greater and three others with levels of 90-99 mSv. Among those born elsewhere, there were five cases (expected 0.74; ratio 6.7, p < 0.01), of which only one was associated with a high level of such radiation. CONCLUSIONS--Paternal preconceptional radiation cannot be the sole cause of the excess in Seascale since it will not explain the excess among those born outside Seascale. It follows that, unless two causes are to be postulated, any single cause must be a factor other than paternal preconceptional radiation. On this basis, the association found among those born there, if not partly due to chance, may reflect an indirect relation with the true cause. The recent hypothesis about such paternal radiation has originated in a subgroup of the excess cases that have aroused concern.     

Can early wilting of old leaves account for much of the ABA accumulation in flooded pea plants?

When pea plants (Pisum sativum L.) were subjected to flooding,abscisic acid (ABA) content in shoots and roots increased upto 8-fold in the following days and stomatal conductance significantlydecreased. Although young leaves of flooded plants had a slightlyhigher water potential than those of the unflooded plants, oldleaves had lower water potential and lost turgor at the timewhen a substantial ABA increase was detected. In plants wherethe old leaves were clipped off, flooding did not cause anyABA increase during 7 d of the experimental period, except underconditions of higher transpiration demand, when the increasein ABA content was both delayed and small in scale (only I-fold).When intact plants were flooded and ABA was assayed separatelyin both old and young leaves, the ABA increase in old leavespreceded that in young leaves. Evidence here suggests that theflooding-induced ABA increase mainly results from the wiltingof old leaves. This suggests that young leaves may be protectedfrom wilting by ABA originating in old leaves under unfavourableenvironmental conditions. Key words: Waterlogging, soil flooding, ABA, leaf water relations, pea, Pisum sativum     

Can changes in sarcolemmal membranes account for the altered inotropic responsiveness in hypertrophied heart?

Hypertrophy is an adaptive mechanism of the heart subjected to pressure overload. Ultrastructural, electrophysiological and mechanical changes occur during this adaptation. A decrease in the inotropic responsiveness of the hypertrophied heart has often been observed as compared to the normal heart. Four sarcolemmal mechanisms that could account for this modification have been described. The mechanism of action of each system (calcium channel, alpha-and beta-adrenergic systems, (Na+,K+)-ATPase) of the hypertrophied heart has been compared to that of the normal heart. In spite of the paucity of results available relating to the calcium channel, the lengthening of the action potential in every case of compensatory hypertrophy could be explained by an altered functioning of the calcium channel. alpha- and beta-adrenergic systems in the hypertrophied heart could be modified at the receptor level itself, or at another level in the cascade of events under their control. For example, two different models of hypertrophy showed a decreased inotropic responsiveness correlated to a defect in the GS regulatory protein. The modification of the ouabain-receptor (Na+,K+)-ATPase mediates a decrease and a prolongation of the inotropic response. According to the modifications of each system, a direct relationship does not seem to exist between the stimulated membrane system and the inotropic responsiveness of the hypertrophied heart.     

Reply to ‘Measurement of nitrogenase activity in legume root nodules: In defense of the acetylene reduction assay’ by J.K. Vessey

This article is in response to that of Vessey (1994) who argues that the traditional, closed acetylene reduction assay can still be a valuable tool for measuring relative differences in nitrogenase activity of legumes. To counter this assertion we consider the practical uses of the traditional assay procedure in relation to real research situations. This requires the use of the assay to be considered separately in the different circumstances of pot-grown and field-grown plants. We conclude that for pot-grown legumes there are a few practical applications where the use of the traditional, closed assay procedure is valid and we accept that these can be extended by the careful use of calibrations against open, flow-through systems. However, we doubt that there are many situations where such a calibration approach would have practical advantages over using the flow-through system to obtain the actual measurements. We cannot recommend any form of the uncalibrated acetylene reduction assay for field-based studies and suggest that researchers consider the merits of simple, alternative measurements such as dry weight, yield and total nitrogen.     

The effect of high pressure homogenization on the activity of a commercial β-galactosidase

High pressure homogenization (HPH) has been proposed as a promising method for changing the activity and stability of enzymes. Therefore, this research studied the activity of β-galactosidase before and after HPH. The enzyme solution at pH values of 6.4, 7.0, and 8.0 was processed at pressures of up to 150?MPa, and the effects of HPH were determined from the residual enzyme activity measured at 5, 30, and 45?°C immediately after homogenization and after 1?day of refrigerated storage. The results indicated that at neutral pH the enzyme remained active at 30?°C (optimum temperature) even after homogenization at pressures of up to 150?MPa. On the contrary, when the β-galactosidase was homogenized at pH 6.4 and 8.0, a gradual loss of activity was observed, reaching a minimum activity (around 30?%) after HPH at 150?MPa and pH 8.0. After storage, only β-galactosidase that underwent HPH at pH 7.0 retained similar activity to the native sample. Thus, HPH did not affect the activity and stability of β-galactosidase only when the process was carried out at neutral pH; for the other conditions, HPH resulted in partial inactivation of the enzyme. Considering the use of β-galactosidase to produce low lactose milk, it was concluded that HPH can be applied with no deleterious effects on enzyme activity.