In Arabidopsis thaliana, trichome cells are specialized unicellular structures with uncertain functions. Based on earlier observations that one of the genes involved in cysteine biosynthesis (Atcys-3A) is highly expressed in trichomes, we have extended our studies in trichome cells to determine their capacity for glutathione (GSH) biosynthesis. First, we have analyzed by in situ hybridization the tissue-specific expression of the genes Atcys-3A and sat5, which encode O-acetylserine(thio)lyase (OASTL) and serine acetyltransferase (SAT), respectively, as well as gsh1 and gsh2, which encode ␥-glutamylcysteine synthetase and glutathione synthetase, respectively. The four genes are highly expressed in leaf trichomes of Arabidopsis, and their mRNA accumulate to high levels. Second, we have directly measured cytoplasmic GSH concentration in intact cells by laser-scanning microscopy after labeling with monochlorobimane as a GSH-specific probe. From these measurements, cytosolic GSH concentrations of 238 ؎ 25, 80 ؎ 2, and 144 ؎ 19 M were estimated for trichome, basement, and epidermal cells, respectively. Taking into account the volume of the cells measured using stereological techniques, the trichomes have a total GSH content more than 300-fold higher than the basement and epidermal cells. Third, after NaCl treatment, GSH biosynthesis is markedly decreased in trichomes. Atcys-3A, sat5, gsh1, and gsh2 mRNA levels show a decrease in transcript abundance, and [GSH] cyt is reduced to 47 ؎ 5 M. These results suggest the important physiological significance of trichome cells related to GSH biosynthesis and their possible role as a sink during detoxification processes.cysteine ͉ glutathione ͉ two-photon laser-scanning microscopy ͉ O-acetylserine(thiol)lyase ͉ sulfate assimilation
Algae and vascular plants are cysteine (Cys) prototrophs. They are able to import, reduce, and assimilate sulfate into Cys, methionine, and other organic sulfur-containing compounds. Characterization of genes encoding the enzymes required for Cys biosynthesis from the unicellular green alga Chlamydomonas reinhardtii reveals that transcriptional and posttranscriptional mechanisms regulate the pathway. The derived amino acid sequences of the C. reinhardtii genes encoding 5Ј-adenylylsulfate (APS) reductase and serine (Ser) acetyltransferase are orthologous to sequences from vascular plants. The Cys biosynthetic pathway of C. reinhardtii is regulated by sulfate availability. The steady-state level of transcripts and activity of ATP sulfurylase, APS reductase, Ser acetyltransferase, and O-acetyl-Ser (thiol) lyase increase when cells are deprived of sulfate. The sac1 mutation, which impairs C. reinhardtii ability to acclimate to sulfur-deficient conditions, prevents the increase in accumulation of the transcripts encoding these enzymes and also prevents the increase in activity of all the enzymes except APS reductase. The sac2 mutation, which does not affect accumulation of APS reductase transcripts, blocks the increase in APS reductase activity. These results suggest that APS reductase activity is regulated posttranscriptionally in a SAC2-dependent process.Plants and algae are primary producers. They absorb sunlight and through photosynthesis, convert it into chemical energy stored as carbohydrate. They also import inorganic nutrients from their environment and convert them into biologically active compounds. Photosynthesis and nutrient acquisition are coordinated with growth.Photosynthetic organisms acclimate to nutrientdeficient conditions through a suite of physiological changes that can be classified as general and specific (Harder and Dijkhuizen, 1983;Davies and Grossman, 1998). General responses, which occur when an organism is deprived of any essential nutrient, include a decrease in the rate of photosynthesis, a decrease in or cessation of cell division, and an accumulation of starch or glycogen. Specific changes occur in response to loss of a specific nutrient and are different for each nutrient. Specific responses are those that enable the organism to scavenge the limiting nutrient from internal or external sources, and those that increase the efficiency of nutrient assimilation.The ability to sense and respond to a nutrientlimiting environments is necessary for an organism to successfully compete in natural environments where nutrients are often limiting. However, the mechanisms used to sense nutrient availability and control physiological changes in response to nutrient limitation are largely unknown. We are using the unicellular green alga Chlamydomonas reinhardtii as a model system to study sulfur metabolism and the response to sulfur-deficient conditions. Sulfur is considered a macronutrient because it is required at relatively high levels. Sulfur is a constituent of proteins, lipids, carbohydrates, el...
Patients admitted to the intensive care unit with criteria of systemic inflammatory response syndrome had a more severe oxidative stress than patients without this syndrome.
Molybdenum is absolutely required for the nitrate-reducing activity of the nicotinamide adenine dinucleotide nitrate reductase complex isolated from Chlorella fusca. The The assimilatory nitrate-reducing system of the green alga Chlorella has been thoroughly characterized in recent years (13-16, 21, 23, 24, 28, 29) and has been shown to be similar to that of higher plants (6,9 (6,8,9,17). However, although molybdenum was early identified by Nicholas and Nason (19) as the metal prosthetic group of nitrate reductase from soybean leaves, workers in the field (3, 6, 14) have until recently been unable to find evidence for either its presence or its function as an electron carrier in nitrate reductase preparations purified from a variety of green plants species. Hewitt and coworkers (1, 2, 9) have reported that, in molybdenum-deficient plants grown in the presence of nitrate, molybdenum is required for the synthesis of nitrate reductase and have suggested that it is involved in the induction process and acts not merely as the constituent metal. Heimer et al. (7) and Wray and Filner (27) have recently studied the effect of tungstate as a competitive inhibitor of molybdate on nitrate assimilation in higher plant tissues and have described the structural and functional relationships of enzyme activities induced by nitrate in barley. Their results support the idea that the nitrate reductase complex is formed in the presence of tungstate but is functional only with respect to NADH-diaphorase. By adding radioactive 9Mo (as molybdate) to a culture of Chlorella cells at the moment derepression of the enzymes of the nitrate-reducing system was initiated by removal of ammonia from the medium, we have recently been able to demonstrate that the metal becomes incorporated into nitrate reductase and remains associated with it during purification (3). Furthermore, we showed that after a mild heat treatment of the enzyme exogenous molybdate chemically reduced by hydrosulfite can be used as electron donor for the enzymatic reduction of nitrate (3).The present work is intended to clarify the role played by molybdenum in nitrate reduction by Chlorella. MATERIALS AND METHODSChlorella fusca Shihira et Kraus (pyrenoidosa, strain 211-15 from Pringsheim's culture collection at Gottingen) was grown autotrophically as described previously (29). When ammonia or ammonium nitrate replaced nitrate as the source of nitrogen, the molarity of each compound was also maintained at 8 mM. In the experiments where ammonium ions were involved, the culture media were buffered with 20 mm sodium phosphate, pH 7.5. In the experiments involving the effect of metals, molybdate was omitted from the standard nutrient solution and either sodium molybdate or sodium tungstate was added as indicated. The algae used for inoculation were grown on nitrate media lacking added molybdate in order to produce a low molybdenum inoculum. Growth was determined by meas-294 www.plantphysiol.org on March 22, 2019 -Published by Downloaded from
The expression of Atcys-3A gene coding for cytosolic O-acetylserine(thiol)lyase, a key enzyme in cysteine biosynthesis, from Arabidopsis thaliana is significantly induced by exposure to salt and heavy-metal stresses. Addition of NaCl to mature plants induced a rapid accumulation of the mRNA throughout the leaf lamina and roots, and later on in stems, being mainly restricted to vascular tissues. The salt-specific regulation of Atcys-3A was also mediated by abscisic acid (ABA) since: (1) exogenous addition of ABA to the culture medium mimicked the salt-induced plant response by raising the level of Atcys-3A transcript, and (2) Arabidopsis mutants aba-1 and abi2-1 were not able to respond to NaCl. Our results suggest that a high rate of cysteine biosynthesis is required in Arabidopsis under salt stress necessary for a plant protection or adaptation mechanism. This hypothesis was supported by the observation that intracellular levels of cysteine and glutathione increased up to 3-fold after salt treatment.
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