共查询到14条相似文献,搜索用时 0 毫秒
1.
Thomas W. Becker Catherine Perrot-Rechenmann Akira Suzuki Bertrand Hirel 《Planta》1993,191(1):129-136
The cellular localization of the enzymes involved in primary nitrogen assimilation was investigated following separation of mesophyll protoplasts and bundle-sheath cells of maize (Zea mays L.) leaves. Determination of the enzymatic activities in the two types of cell revealed that nitrate and nitrite reductase are principally located in the mesophyll cells whereas glutamine synthetase (GS) and ferredoxin-dependent glutamate synthase (Fd-GOGAT) are present in both tissues with a preferential location in the bundle-sheath strands. In order to confirm the results obtained by this conventional biochemical method we have used an in-situ immunofluorescence technique to unambiguously localize GS and Fd-GOGAT at the cellular level. Thin-sectioned maize leaves treated with specific GS and Fd-GOGAT antisera followed by conjugation with fluorescein-isothiocyanate-labelled sheep anti-rabbit immunoglobulins clearly show that GS is equally distributed within the leaf whereas Fd-GOGAT is mostly present in the chloroplasts of the bundle-sheath cells. The cellular localization of nitrate reductase, nitrite reductase, GS-2 and Fd-GOGAT in maize leaf cell types strongly indicates that primary nitrogen assimilation functions in the mesophyll cells while photorespiratory nitrogen recycling is restricted to the bundle-sheath cells. 相似文献
2.
The activities of nitrate reductase (EC1.6.6.1), nitrite reductase (EC 1.6.6.4), glutamine synthetase (EC6.3.1.2), glutamate synthase (EC1.4.7.1) and NAD(P)H-dependent glutamate dehydrogenase (EC 1.4.1.3) were investigated in mesophyll and bundle sheath cells of maize leaves (Zea mays L.). Whereas nitrate and nitrite reductase appear to be restricted to the mesophyll and GDH to the bundle sheath, glutamine synthetase and glutamate synthase are active in both tissues.During the greening process, the activities of nitrate and nitrite reductase increased markedly, but glutamine synthetase, glutamate synthase and glutamate dehydrogenase changed little.Abbreviations BDH
British Drug Houses
- EDTA
Ethylene diamine tetra-acetic acid
- GDH
Glutamate dehydrogenase
- NADH
Nicotinamide-adenine dinucleotide reduced form
- NADPH
Nicotnamide-adenine dinucleotide phosphate reduced form
- PMSF
Phenylmethyl sulphonyl fluoride 相似文献
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Summary. Accumulation of amino acids was studied in rice roots of 3-day-old seedlings subjected for 48 h to anaerobic conditions.
Alanine and Gaba were the main amino acids accumulated under anoxia. Their synthesis was strongly inhibited by MSX and AZA,
inhibitors of glutamine synthetase and glutamate synthase. These activities increased after 8 h of anaerobic treatment and,
by immunoprecipitation of 35S-labeled proteins, it was shown that glutamine synthetase and ferredoxin-dependent glutamate synthase were synthesized during
the treatment. These findings indicate that the glutamine synthetase/glutamate synthase cycle play an important role in anaerobic
amino acid accumulation.
Received April 5, 1999 相似文献
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Glutamine synthetase from mesophyll and bundle sheath maize cells: isoenzyme complements and different sensitivities to phosphinothricin 总被引:13,自引:0,他引:13
B. González-Moro A. Mena-Petite M. Lacuesta C. González-Murua A. Muñoz-Rueda 《Plant cell reports》2000,19(11):1127-1134
Anion-exchange FPLC has been used to resolve the isoforms of glutamine synthetase (GS, EC 6.3.1.2) from Zea mays mesophyll (MC) and bundle sheath cells (BSC). Two different isoforms were detected in both types of photosynthetic cells.
The predominantly active isoform was GS1 (61%) in MC and GS2 (67%) in BSC. The relative contribution of GS1 and GS2 to the
overall GS activity in BSC in maize here reported resembles the proportion described for most C3 plants. Differences among
these isoforms in terms of their susceptibility to phosphinothricin (PPT), an analogue of glutamate and known inhibitor of
GS, were found. The GS1 isoenzyme from MC was the most sensitive form, being inhibited by 50% at approximately 2.0 μM DL-PPT, whereas the GS2 from BSC presented the highest tolerance to the inhibitor (I50=30 μM). The transferase-to-semibiosynthetic activity ratio for the MC isoforms, which was higher than the ratio for the BSC isoforms,
and the differences shown by the isoforms in susceptibility to PPT predict important differences in the biochemical properties
and regulation of GS isoenzymes. In this regard, the cytoplasmic isoenzymes, and especially the one in MC, due to its relatively
high contribution to mesophyll cell GS activity, could play a vital role in nitrogen metabolism in maize.
Received: 1 December 1999 / Revised: 7 February 2000 / Accepted: 23 February 2000 相似文献
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Different pH-dependences of K+ channel activity in bundle sheath and mesophyll cells of maize leaves
The isolation of bundle sheath protoplasts from leaves of Zea mays L. for patch clamp whole-cell experiments presents special problems caused by the suberin layer surrounding these cells.
These problems were overcome by the isolation technique described here. Two different types of whole-cell response were found:
a small response caused by MB-1 (maize bundle sheath conductance type 1) which was instantaneously activated, and another
caused by MB-2 (maize bundle sheath conductance type 2) consisting of an instantaneous response (maize bundle sheath K+ instantaneous current type 2; MB-KI2) similar to but stronger than the current through MB-1 plus a small time-dependent outward
rectifying component (maize bundle sheath activated outward rectifying current; MB-AOR) with voltage-dependent delayed activation.
The occurrence of MB-AOR was often accompanied by a smaller contribution from an inward rectifying channel at negative potentials.
Activation of MB-2 required ATP. It is suggested that MB-1 and MB-2 are related to bundle sheath cells with and without direct
contact with the xylem vessels. In mesophyll cells, only one type of response caused by MM-2 (maize mesophyll conductance
type 2) was found with an instantaneous (maize mesophyll K+ instantaneous current type 2, MM-KI2) and a voltage-dependent delayed component (maize mesophyll activated outward rectifying
current, MM-AOR). The most striking difference between bundle sheath and mesophyll cells was the pH dependence of K+ uptake. At pH 7.2, uptake of K+ by MB-2 was identical to that by MM-2 over the whole voltage range. However, acidification stimulated K+ conductance in bundle sheath cells, whereas a decrease was found for MM-2. At pH 6.15, the bundle sheath channel MB-2 had
more than a 10-fold higher K+ uptake at positive and negative potentials than MM-2. The channel MB-1, too, was stimulated by low pH. This seems to indicate
a putative role for MB-1 and MB-2 in charge balance during uptake of nutrients via cotransport from the xylem into the symplasm.
Received: 23 April 1999 / Accepted: 19 July 1999 相似文献
10.
In malic enzyme-dependent crassulacean-acid-metabolism (ME-CAM) plants, malic acid is decarboxylated by NADP-ME and NAD-ME and generates pyruvate with CO2. Pyruvate is phosphorylated to phosphoenolpyruvate by pyruvate, Pi dikinase (PPDK) and is then conserved in gluconeogenesis. Although PPDK was considered to be located in chloroplasts (e.g., Mesembryanthemum crystallinum), it has recently been found to accumulate in both the chloroplasts and the cytosol in two Kalancho? species. In this study, the intracellular localization of PPDK was investigated in 22 ME-CAM species in 13 genera of 5 families by immunogold labeling and electron microscopy. This revealed that the pattern of intracellular localization of PPDK varies among the ME-CAM plants and is divided into three types: Chlt, in which PPDK accumulates only in the chloroplasts; Cyt-Chlt, in which PPDK accumulates in both chloroplasts and cytosol; and Cyt, in which PPDK accumulates predominantly in the cytosol. Members of a particular genus tend to have a common PPDK-localization type. In the Cactaceae, all species from seven genera were classified as Cyt. The photosynthetic tissues of all ME-CAM species, including the Cyt type, had substantial PPDK activity, suggesting that PPDK in the cytosol is active and probably plays a functional role. In the Chlt species, NADP-ME activity was relatively greater than NAD-ME activity. In the Cyt-Chlt and Cyt species, however, either the activity of NAD-ME was higher than that of NADP-ME or they were approximately the same. The species variation in the intracellular localization of PPDK is discussed in relation to CAM function and to molecular and phylogenetic aspects. 相似文献
11.
Dick B. Janssen Patricia M. Herst Han M. L. J. Joosten Chris van der Drift 《Archives of microbiology》1981,128(4):398-402
In Pseudomonas aeruginosa the formation of urease, histidase and some other enzymes involved in nitrogen assimilation is repressed by ammonia in the growth medium. The key metabolite in this process appears to be glutamine or a product derived from it, since ammonia and glutamate did not repress urease and histidase synthesis in a mutant lacking glutamine synthetase activity when growth was limited for glutamine. The synthesis of these enzymes was repressed in cells growing in the presence of excess glutamine. High levels of glutamine were also required for the derepression of NADP-dependent glutamate dehydrogenase formation in the glutamine synthetase-negative mutant. 相似文献
12.
Infiltrating detached maize (Zeamays L.) leaves with L-galactono-1,4-lactone (L-GAL) resulted in a 4-fold increase in the content of leaf ascorbate. Upon exposure to high irradiance (1000 μmol photons m−2 s−1) at 5 °C, L-GAL leaves de-epoxidized the xanthophyll-cycle pigments faster than the control leaves; the maximal ratio of de-epoxidized
xanthophyll-cycle pigments to the whole xanthophyll-cycle pool was the same in both leaf types. The elevated ascorbate content,
together with the faster violaxanthin de-epoxidation, did not affect the degree of photoinhibition and the kinetics of the
recovery from photoinhibition, assayed by monitoring the maximum quantum efficiency of photosystem II primary photochemistry
(Fv/Fm). Under the experimental conditions, the thermal energy dissipation seems to be zeaxanthin-independent since, in contrast
to the de-epoxidation, the decrease in the efficiency of excitation-energy capture by open photosystem II reaction centers (Fv′/Fm′) during the high-irradiance treatment at low temperature showed the same kinetic in both leaf types. This was also observed
for the recovery of the maximal fluorescence after stress. Furthermore, the elevated ascorbate content did not diminish the
degradation of pigments or α-tocopherol when leaves were exposed for up to 24 h to high irradiance at low temperature. Moreover,
a higher content of ascorbate appeared to increase the requirement for reduced glutathione.
Received: 20 May 1999 / Accepted: 29 October 1999 相似文献
13.
Non-photochemical quenching of chlorophyll fluorescence (NPQ) and quantum yield of photosystem II (PSII) were studied with
intact mesophyll chloroplasts of maize (Zea mays L.) during the initial minutes of illumination using the pulse-modulated chlorophyll fluorescence technique. Non-photochemical
quenching was rapidly reversible in the dark at any point during illumination, which is indicative of energy-dependent dissipation
of energy (mediated via thylakoid ΔpH changes and ascorbate-dependent synthesis of zeaxanthin). In chloroplasts suspensions
including 15 mM ascorbate in the medium, with addition of oxaloacetate and pyruvate, the PSII yield, rate of reduction of
oxaloacetate and phosphorylation of pyruvate reached a maximum after approximately 2 min of illumination. Under these conditions,
which promote phosphorylation and a decreased ΔpH across the thylakoid membrane, NPQ rose to a maximum after 2–3 min of illumination,
dropped to a minimum after about 6 min, and then increased to a steady-state level. A rather similar pattern was observed
when leaves were illuminated following a 30-min dark period. Providing chloroplasts with higher levels of ascorbate (60 mM),
prevented the transient drop in NPQ. Anaerobic conditions or addition of potassium cyanide caused a decrease in PSII yield,
providing evidence for operation of the ascorbate-dependent Mehler-peroxidase reaction. These conditions also strongly suppressed
the transient drop in NPQ. Dithiothreitol, an inhibitor of violaxanthin de-epoxidase, caused a large drop in NPQ even in the
presence of high levels of ascorbate. The results suggest that the decline of NPQ occurs in response to an increase in lumen
pH after initiation of phosphorylation, that this decline can be suppressed by conditions where ascorbate is not limiting
for violaxanthin de-epoxidase, and that the increase of NPQ after such a decline is the result of development of energy dissipation
in PSII reaction centers.
Received: 13 August 1999 / Accepted: 17 September 1999 相似文献
14.
No active uptake of ammonium was detected in Proteus vulgaris, Bacillus pasteurii, and Sporosarcina ureae, which indicates that these bacteria depend on the passive diffusion of ammonia across the cell membrane. In P. vulgaris the glutamine synthetase-glutamate synthase (GS-GOGAT) pathway and glutamate dehydrogenase (GDH) were present, and these enzymes exhibited high affinities for ammonium. In B. pasteurii and S. ureae, however, no GS activity was detected, and GOGAT activity was only present in S. ureae. GDH enzymes were present in these two organisms, but showed only low affinity for ammonium, with apparent K
m-values of 55.2 mM in B. pasteurii and 36.7 mM in S. ureae, repectively. These observations explain why P. vulgaris is able to grow at neutral pH and low ammonium concentration (2 mM), while B. pasteurii and S. ureae require high ammonium concentration (40 mM) and alkaline pH for growth.Non-standard abbreviations GS
glutamine synthetase
- GOGAT
glutamate synthase
- GDH
glutamate dehydrogenase
- GT
glutamyl transferase
- MA
methylammonium
- NB
nutrient broth
- YE
yeast extract
- NA
nocotinic acid 相似文献