Induction of Glutamine Synthetase During Nodule Development in Lupin

Robertson, John; Farnden, Kevin J. F.; Warburton, Marion P.; Banks, J. A. M.

doi:10.1071/pp9750265

Cited by 89 publications

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“…That leghaemoglobin synthesis precedes nitrogenase activity is consistent with the results of Godfrey et al ( 1 975) (R. Zupini, leghaemoglobin determined immunologically), Robertson et al (1975) …”

Section: Discussionsupporting

confidence: 81%

The Sequence of Appearance of Leghaemoglobin and Nitrogenase Components I and II in Root Nodules of Pisum sativum

et al. 1980

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~~~ ~ ~~The sequence of appearance of nitrogenase components I and TI and leghaemoglobin in root nodules of pea plants inoculated with Rhizobium Zeguminosarum (strain PRE) was studied using radioimmunoassays. Leghaemoglobin was detected before nitrogenase activity. The MoFe component (I) of nitrogenase was detected at the same time as leghaemoglobin, while the Fe component (11) of nitrogenase and nitrogenase activity appeared 1 and 2 d later, respectively. The same order of appearance of nitrogenase activity and leghaemoglobin was found in root nodules of cowpea plants inoculated with Rhizobium sp. 32H 1. I N T R O D U C T I O NIn an earlier paper (Bisseling et al., 1979) we demonstrated that leghaemoglobin in pea root nodules induced by Rhizobium Zeguminosarzm was synthesized before nitrogenase activity could be measured. We also found that in the early phase of the development of nitrogen-fixing activity, the synthesis of the MoFe component (CI) of nitrogenase exceeded the synthesis of the Fe component (CII) (Bisseling et al., 1979). In this paper we report further experiments on the order of synthesis of leghaemoglobin and CI and CII of nitrogenase, which are three essential components for nitrogen fixation in pea-rhizobium symbiosis. A radioimmunoassay has been developed to quantify leghaemoglobin and C1 and CIJ. This method is a few orders of magnitude more sensitive than the Mancini immunodiffusion assay (Mancini et al., 1965), which we used previously for the determination of leghaemoglobin; it has allowed us to extend the measurements towards the beginning of nodule development and to confirm that leghaemoglobin is present before nitrogenase activity is measurable. Moreover, we have now found that CI is synthesized before the presence of CII can be demonstrated in bacteroid extracts.Fast-and slow-growing rhizobia differ in many respects (e.g. Paau, 1978). To investigate whether slow-growing rhizobium bacteroids differ from the fast-growing rhizobia (R.legurninusarum) in the order of appearance of nitrogenase activity and leghaemoglobin synthesis, we have studied the appearance of leghaemoglobin and nitrogenase activity in root nodules of cowpea plants inoculated with Rhizobium sp. 32Hl. The results were similar to those obtained with the pea root nodules. METHODS Growth of plants.Pea plants (Pisum sativunz var. Rondo) were cultured and nodulated with Rhizobium ieguminosarum (strain PRE), as described earlier (Bisseling et al., 1978), at 19 to 20 "C with an 8 h dark and 16 h light period. Rhizobium leguminosarum was cultured in a yeast/mannitol medium (Bisseling et al., 1978). Vigna unguiculata T . B I S S E L I N G A N D O T H E R SAssay of nitrogenase. Nitrogenase activity of intact nodules was measured by the acetylene reduction method on pieces of the main root carrying the root nodules. Ethylene concentrations were measured in a Pye 104 gas chromatograph with a Porapak R column (Bisseling et al., 1978).Preparation of bacteroid and plant proteins. The isolation of bacteroids and the preparation of s...

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Section: Discussionsupporting

confidence: 81%

The Sequence of Appearance of Leghaemoglobin and Nitrogenase Components I and II in Root Nodules of Pisum sativum

et al. 1980

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“…Soluble protein of each supernatant fraction was determined by the method of Lowry et al (18) using BSA, fraction V, as the protein standard. GS activity was determined according to Robertson et aL (25), but because higher rates were observed with 50 mm Hepes buffer (pH 7.0) it was used instead of the tris(hydroxymethyl)aminomethane-maleic acid buffer. Duplicate assays were made for each sample.…”

Section: Methodsmentioning

confidence: 99%

Ontogenetic Variation of Nitrogenase, Nitrate Reductase, and Glutamine Synthetase Activities in Oryza sativa

Berkum

Sloger²

1981

Plant Physiol.

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The relatship between the rates of nitrogenase, nitrate reductase, and gluta e synthetase acdviies, and plant ontogeny in rice (O'yza sativa L.), cultivar 'M9', grown in salt marsh sediment with and without nitrate treatment was studied. In both treatments, nitrogenase activity measured as the Immediate lnear rate of acetylene reduction by bacteria associated with the roots varied with plant age. In control plants, the nitrogenase activity devel During the vegetative and early reproductive stages of growth, the development of maximal rates of nitrogenase activity coincided with an increase of total nitrogen of the plants in both treatments.Grasses are considered to assimilate mainly inorganic nitrogen for growth, but they may stimulate nitrogen fixation by bacteria associated with their roots (6, 9, 10). The rate of nitrogen fixation in legumes has been shown to be dependent on plant age (13). The seasonal variations of leaf NRA3 and nitrogen fixation in soybeans have been reported, and each contributes nitrogen during plant development (14, 28 3 Abbreviations: NRA, nitrate reductase activity; GS, glutamine synthetase; GOGAT, glutamine:a-ketoglutarate aminotransferase.(20).The rate of nitrogenase activity associated with grass roots has been reported also to vary with plant development (2,7,8,17,23,24). The rates of leaf NRA and nitrogenase activity have been suggested to peak successively (2, 7, 24). However, the measurements of acetylene reduction were made with excised roots after an overnight incubation. This procedure was adopted because acetylene reduction by excised grass roots was not detected for 8 to 18 h (1). Acetylene reduction by excised grass roots after a long delay is caused by the induction of nitrogenase activity in a proliferating microbial population (3). In a recent review, van Berkum and Bohlool (6) concluded that acetylene reduction by excised roots after an extended delay does not indicate the ability of grasses to support and benefit from nitrogen fixation. In contrast, excised nodulated legume roots reduce acetylene immediately at linear rates (13), and the reported rates of nitrogenase activity in legumes are based on short term assays.We have reported immediate linear rates of acetylene reduction by several salt marsh grasses and rice grown in salt marsh sediment (5). The immediate linear rates of acetylene reduction by rice roots removed from the sediment are consistent with the kinetics of the detection of nitrogenase activity by other nitrogen-fixing systems. Therefore, rice roots have the ability to support nitrogen fixation by associated bacteria. Preliminary studies of 15N2 incorporation have demonstrated that rice may benefit from root-and leafassociated nitrogen fixation (15).The objectives of this work were: (a) to determine how the rate of root-associated nitrogenase activity in rice varies with plant ontogeny using techniques that measure immediate linear rates of acetylene reduction and (b) to study the relationship between the rates of nitrogenase, NRA, and GS a...

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“…The pattern of expression of the various cytosolic GSs has been most thoroughly investigated in legumes where GS is actively synthesized in root nodules to assimilate the large supply of ammonium produced by the symbiotic N-fixing bacteroids (Robertson et al, 1975;Lara et al, 1983). Although different from one legume species to the other (temperate legumes versus tropical legumes forming either determinate or indeterminate nodules), it has been shown clearly that metabolic and/or developmental events control the expression of some members of the nodule GS multigene family in a tissue-specific manner.…”

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confidence: 99%

Does Lowering Glutamine Synthetase Activity in Nodules Modify Nitrogen Metabolism and Growth of Lotus japonicus?

et al. 2003

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A cDNA encoding cytosolic glutamine synthetase (GS) from Lotus japonicus was fused in the antisense orientation relative to the nodule-specific LBC3 promoter of soybean (Glycine max) and introduced into L. japonicus via transformation with Agrobacterium tumefaciens. Among the 12 independent transformed lines into which the construct was introduced, some of them showed diminished levels of GS1 mRNA and lower levels of GS activity. Three of these lines were selected and their T 1 progeny was further analyzed both for plant biomass production and carbon and nitrogen (N) metabolites content under symbiotic N-fixing conditions. Analysis of these plants revealed an increase in fresh weight in nodules, roots and shoots. The reduction in GS activity was found to correlate with an increase in amino acid content of the nodules, which was primarily due to an increase in asparagine content. Thus, this study supports the hypothesis that when GS becomes limiting, other enzymes (e.g. asparagine synthetase) that have the capacity to assimilate ammonium may be important in controlling the flux of reduced N in temperate legumes such as L. japonicus. Whether these alternative metabolic pathways are important in the control of plant biomass production still remains to be fully elucidated.Nitrogen (N) is one of the major limiting factors for plant growth. However, an excessive external supply of N causes major problems in agriculture and the environment, mostly due to nitrate leaching into underground water (Benes et al., 1989), not only polluting the aquatic environment but also resulting in a high nitrate content in food and drinks (Moller et al., 1990). Paradoxically, dinitrogen is 80% of the atmosphere, but it can only be assimilated by prokaryotic symbiotic or free-living diazotrophic organisms possessing nitrogenase activity (Dilworth, 1974). In higher plants, nitrate and ammonium are the two major inorganic N compounds that can be directly assimilated (Beevers, 1976). They are provided either artificially by the external supply of fertilizers, bacterial nitrification, or atmospheric reduction of dinitrogen during lightning, or naturally through biological N fixation. Therefore, regulation of inorganic N assimilation and incorporation of inorganic N into organic matter is of major importance not only in maintaining a sustainable agriculture but also protecting the environment.Over the past few years, attention has been particularly focused on the enzyme Gln synthetase (GS; E.C. 6.3.1.2) because of its central role in N metabolism and its diverse metabolic and developmental regulation in different plant species and organs (Cren and Hirel, 1999). Two major isoforms exist for the GS enzyme: cytosolic GS, occurring in the cytoplasm of leaves and non photosynthetic organs, and chloroplastic GS, present only in the chloroplasts of photosynthetic tissues and the plastids of roots or etiolated plants. The relative proportions of the cytosolic and plastidial GS activity may vary within different organs of the same plant or within dif...

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Induction of Glutamine Synthetase During Nodule Development in Lupin

Cited by 89 publications

References 0 publications

The Sequence of Appearance of Leghaemoglobin and Nitrogenase Components I and II in Root Nodules of Pisum sativum

The Sequence of Appearance of Leghaemoglobin and Nitrogenase Components I and II in Root Nodules of Pisum sativum

Ontogenetic Variation of Nitrogenase, Nitrate Reductase, and Glutamine Synthetase Activities in Oryza sativa

Does Lowering Glutamine Synthetase Activity in Nodules Modify Nitrogen Metabolism and Growth of Lotus japonicus?

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