JXB Advance Access originally published online on December 20, 2004
Journal of Experimental Botany 2005 56(412):695-702; doi:10.1093/jxb/eri054
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
RESEARCH PAPER |
Accumulation and remobilization of amino acids during senescence of detached and attached leaves: in planta analysis of tryptophan levels by recombinant luminescent bacteria
1Faculty of Biology, Technion, Israel Institute of Technology, Haifa 32000, Israel
2Faculty of Biotechnology and Food Engineering, Technion, Haifa 32000, Israel
* To whom correspondence should be addressed. Fax: +972 4 8225153. E-mail: gepstein{at}tx.technion.ac.il
Received 21 June 2004; Accepted 21 October 2004
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and methodsResults and discussionReferences |
|---|
The process of leaf senescence is biochemically characterized by the transition from nutrient assimilation to nutrient remobilization. The nutrient drain by developing vegetative and reproductive structures has been implicated in senescence induction. The steady-state levels of amino acids in senescing leaves are dependent on the rate of their release during protein degradation and on the rate of efflux into growing structures. To determine the possible regulatory role of amino acid content in leaf senescence, an in planta non-destructive, semi-quantitative method for the analysis of endogenous levels of free amino acids has been developed. The method is based on in vivo bioluminescence of amino acid-requiring strains of recombinant Escherichia coli carrying the lux gene. The luminescence signal was found to be proportional to the levels of added exogenous tryptophan and to the free amino acid levels in the plant tissues analysed. During the senescence of tobacco flowers and of detached leaves of oats and Arabidopsis, a progressive increase in the levels of free amino acids was monitored. By contrast to the detached leaves, the attached oat leaves displayed a decrease in the levels of free amino acids during senescence. In Arabidopsis, both the attached and detached leaves exhibited a similar pattern of gradual increase in amino acid content during senescence. The differences between the sink–source balance of the two species and the possible relationships between amino acid content and leaf senescence are discussed.
Key words: Amino acids, Arabidopsis, leaf senescence, luminescent bacteria, lux system, nitrogen, sink, source
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and methodsResults and discussionReferences |
|---|
Senescence is the last stage of leaf development, preceding death, and is characterized by the transition from anabolic to catabolic processes. The chloroplast is the first organelle to show drastic changes, due to the differential breakdown of its components (Gepstein, 1988
). Consequently, a decline in photosynthetic activity occurs that may trigger the induction of the senescence syndrome (Okada and Katoh, 1998). This metabolic turning point is also associated with the transition from nutrient assimilation to nutrient remobilization (Feller and Fisher, 1994; Masclaux et al., 2000). The syndrome of leaf senescence is genetically controlled and normally occurs even under optimal conditions. Recent genomic studies have identified a large number of genes, designated senescence associated genes (SAGs), whose products are likely to initiate, regulate, and participate in the execution phase of the senescence syndrome (Quirino et al., 1999; Dangl et al., 2000; He and Gan, 2002; Bhalerao et al., 2003; Buchanan-Wollaston et al., 2003; Gepstein et al., 2003; Yoshida, 2003; Gepstein, 2004; Guo et al., 2004). Among the prominent SAGs are genes whose products encode hydrolytic enzymes. These enzymes are responsible for the intensive breakdown of the various macromolecules such as proteins, nucleic acids, polysaccharides, and lipids (Nam, 1997; Lers et al., 1998; Thompson et al., 1998; Rubinstein, 2000; Bhalerao et al., 2003; Buchanan-Wollaston et al., 2003; Gepstein et al., 2003; Yoshida, 2003; Guo et al., 2004). The nutrients released by the hydrolytic enzymes are remobilized to growing regions such as young leaves or reproductive tissues. Nitrogen is primarily released from protein breakdown and nucleic acid metabolism (Hortensteiner and Feller, 2002), and was estimated to be the most recycled nutrient (90%) during senescence (Himelblau and Amasino, 2001). Several transporters responsible for intracellular nitrogen translocation have been reported; among them are oligopeptide transporters, aromatic amino acid permeases, ammonium transporter, and purine and pyrimidine transporters (Guo et al., 2004). A positive correlation between the increase in leaf proteolytic activity and the expression of two senescence marker genes that encode glutamine synthetase and glutamate dehydrogenase has been suggested (Terce-Laforgue et al., 2004).
Altering the source–sink balance in the whole plant may affect both the timing and the rate of progression of leaf senescence (Nooden, 1980). In the most extreme demonstration of the so-called correlative control phenomenon, where reproductive organs representing the sink structures were totally removed, either individual leaf or the whole plant senescence was delayed (Nooden, 1980). When the reproductive sink was reduced in transgenic plants with low activity of sucrose phosphate synthase, N mobilization was suppressed (Ono et al., 1999).
Development of reproductive structures does not always affect leaf senescence, and Arabidopsis is one of the exceptions (Hensel et al., 1994; Nooden and Penney, 2001). The longevity of individual rosette leaves is not controlled by reproductive structures, but the longevity of the whole Arabidopsis plant is under correlative control of the reproductive organs (Nooden and Penney, 2001).
Several studies in the past few years have raised the possibility that various metabolites released during the onset of senescence have regulatory roles in the expression of genes or proteins involved in the mobilization of nitrogen, carbon, and minerals later in senescence (Yoshida, 2003; van Doorn, 2004). Prerequisite for these studies is the availability of method(s) to monitor the levels and spatial distribution of metabolites in the various organs and/or tissues. Biochemical analysis of plant metabolites and enzyme activities is usually performed following tissue homogenization, which of course does not necessarily reflect the in vivo metabolite levels or their spatial distribution. It may well be that a spatial separation of the actual compartments and conditions in the living cell prevents various interactions that under in vitro conditions do occur, thus altering the results of metabolite analysis. For macromolecules such as RNA and proteins, in situ and immunolocalization methods have been developed, significantly contributing to the understanding of spatial distribution of gene and protein expression in various biological systems. Similar in situ approaches for the analysis of most metabolites, however, are not yet available.
In the context of leaf senescence, it has been hypothesized that metabolites such as carbohydrates and amino acids may have a role in the induction and in the advance of the senescence programme (Yoshida, 2003). However, to establish this causal relationship between levels of metabolites such as amino acids and the senescence programme, suitable methods for in planta analyses of the metabolites are required.
This paper is a report on the development of an in planta method for analysing endogenous amino acid levels. The suggested method is based on the application of a non-destructive and sensitive bacterial luminescence technology. The luminescence signal is proportional to the amino acid levels in the plant tissues.
The use of bacterial luminescence for various analytical applications has become very popular during the last decade. Most of the described methods are associated with cell-free extracts but, only recently, intact luminous bacteria were found suitable and sensitive for different analytical purposes (Ulitzur, 1997). Here, for the first time, the intact luminous bacteria system is employed for in planta non-destructive semi-quantitative biochemical analysis of amino acids in plant tissues.
Luminous bacteria contain the luciferase enzyme (lux) which simultaneously oxidizes both reduced flavin-mononucleotide (FMNH2) and a long-chain aldehyde to give FMN, water, the corresponding fatty acid, and the emission of blue-green (490 nm) light. Levels of in vivo bacterial luminescence are especially high, and emission by even a single bacterium (5x10 quanta s–1 cell–1) can be readily detected with a photon counter or by image recording. Recombinant strains of Escherichia coli expressing the lux system have been produced and successfully applied for various analytical purposes (Ulitzur, 1997). In vivo bioluminescence of recombinant strains of E. coli carrying the lux gene showed an excellent correlation to bacterial viability. Thus, levels of any promotive or inhibitory factor that affects bacterial viability can be indirectly quantified by the bioluminescence assays. Among the known analytical applications are: determinations of long-chain fatty acids, lipopolysaccharides, ethanol and NAD, NADH, antibiotic activity, toxicant levels, etc. In the context of amino acid analysis, viability and the resulting in vivo bioluminescence of amino acid-requiring mutants of E. coli carrying the lux system depend on the availability of the required amino acid. Linear relationships have been found between increasing levels of the supplied amino acid and the bioluminescence emission of the auxotrophic mutant of E. coli carrying the lux system (Ulitzur, 1997).
In this study, in planta analysis of amino acids has been employed to elucidate the relationships between sink–source interaction and organ senescence. The possible relationships between free amino acid levels and the induction of senescence have been assessed here in two senescence systems: the natural senescence of attached leaves and the senescence of detached leaves. It was also important to determine the degree of universality of the correlative relationships between free amino acid levels and leaf senescence and the possible regulatory role of amino acids in inducing the senescence programme. With this aim, the temporal amino acid patterns of two different monocarpic species, oat and Arabidopsis, were compared.
|
Materials and methods |
|---|
|
TopAbstractIntroductionMaterials and methodsResults and discussionReferences |
|---|
Plant material
Oat (Avena sativa L. cv. Victory) seedlings were grown on soil in a controlled-environment growth chamber (16 h light, 24 °C). For the detached leaf analysis, primary leaves were harvested from 10-d-old plants and were then transferred and incubated on wet Whatman no. 1 filter paper in Petri dishes at 24 °C in darkness. Leaf segments were incubated for the indicated periods (0, 1, 2, 3, 4 d) and thereafter were stored at –80 °C. The attached leaf analysis was performed with primary leaves at different stages during the progression of senescence. Prior to the analysis, leaves were kept at –80 °C.
Arabidopsis thaliana L. cv. Columbia) seedlings were grown in a controlled-environment growth room at 23 °C under 14 h light. For the detached leaf assay, 3 weeks after germination, before the emergence of the inflorescence stem, the 5th and 6th rosette leaves were harvested and incubated at 24 °C on wet Whatman no. 1 filter paper in Petri dishes for the indicated periods in darkness. For the attached leaf assay, the 5th and the 6th rosette leaves were harvested from plants at different ages. Leaves were stored at –80 °C.
Tobacco (Nicotiana tabacum L. cv. SR1) plants were grown on soil in the greenhouse at 25 °C. Flowers at different developmental and senescence stages were harvested and immediately transferred to –80 °C.
Crude extract assays
For analysis of the free amino acid in the plant crude extracts, samples of 200 mg fresh weight of the plant material were homogenized in 1 ml of 0.1 M TRIS-HCl, pH 7.5. Protease inhibitor cocktail for plants (Sigma) was added to the homogenate.
Bacteria and growth conditions
Recombinant tryptophan auxotrophic mutant of E. coli (TK4) (kindly provided from Dr J Kuhn, Department of Biology, Technion) was transformed with the plasmid PW21A (Adar et al., 1992) Formation of the LuxR protein in the Vibrio fischeri lux system is controlled by HtpR through the GroESL proteins carrying the lux system of Vibrio fischeri (luxCDABE) in the absence of the luxI gene. The culture was grown overnight at 37 °C with shaking (250 rpm) in Luria broth medium supplemented with tryptophan (10 µg l–1). The culture was washed three times with saline-phosphate buffer and added at a final concentration of 0.1 OD (600 nm) to the assay medium consisting of Vogel medium at pH 7.0 containing 1.0% agar, 0.1% Difco casamino acids, 4% BSA, and 0.1 µg ml–1 of the V. fischeri inducer N-3-oxohexanoyl-L-homoserine lactone (Sigma).
Bioluminescence assay for amino acids
Leaves or flowers were kept at –80 °C, thawed and placed on the bacterial assay medium at 28 °C. To ensure that analysis was even along the leaf segment, full contact between the whole leaf surface and the assay medium was required. For comparison, plant crude extracts were allowed to soak into 1-cm-diameter discs of Whatman no. 3 filter papers and were then incubated on the agar-containing bacterial assay medium. Following incubation for 1 h, the Petri dishes were transferred into a dark room and were either photographed with a digital camera or exposed to X-OMAT LS Kodak film. The bioluminescence emission was visualized following film development.
Spectrophotometric assays for chlorophyll and total free amino acids
Eight leaf segments incubated in Petri dishes in the dark on Whatman no. 1 filter paper, soaked with 10 ml water, were analysed for their chlorophyll and amino acid content. The extraction of the chlorophyll and amino acids of four leaf segments was carried out in 10 ml of 80% boiled ethanol for 5 min. In addition, amino acid leakage into the incubation medium was determined periodically by analysing 1 ml of the medium for its amino acid content in parallel to the analysis of the endogenous levels (Gepstein and Thimann, 1981). Chlorophyll concentration was spectrophotometrically determined by measuring the absorbance at 645 nm. Aliquots (0.1 ml) from the alcoholic leaf extracts and 1 ml of the corresponding incubation medium were used for amino acid determination by the ninhydrin colorimetric method using leucine as a standard (Gepstein and Thimann, 1981).
|
Results and discussion |
|---|
|
TopAbstractIntroductionMaterials and methodsResults and discussionReferences |
|---|
A bioluminescence in planta analysis of free amino acids
Auxotrophic mutants of E. coli carrying the V. fischeri lux system, proven suitable for detection of various metabolites/growth factors, were evaluated as a detection modality system for in planta analysis of total free amino acids. The luminescence response of the tryptophan-requiring bacteria, which reflects the metabolic activity and viability of the luminescence bacteria, is proportional to the tryptophan levels (between 20 and 100 µg ml–1) added to the bacterial growth media (Fig. 1A). The sensitivity of the luminescence assay also depends on the incubation time and the exposure time of the X-ray film. The physiological level of tryptophan in green leaves is relatively low (5–10 µg g–1) but can still be detected by the luminescence assay (Tozawa et al., 2001
). However, the dramatic increase in the levels of free tryptophan released by the characteristic senescence-associated proteolysis can be easily detected by this method (Fig. 1). During leaf senescence, some of the amino acids are modified before being transported and the amids glutamine and asperagine are the most prominent examples. Tryptophan has not been reported to be modified during leaf senescence. Thus, the changes in the levels of free tryptophan during senescence may also represent the changes in the levels of free amino acids released during the senescence-associated proteolysis. Indeed, a similar pattern of changes in the levels of total free amino acids during leaf senescence was detected by the classical analytical method using ninhydrin (Fig. 2). This similarity further supports the applicability of the bioluminescence method for determining free amino acid levels in senescing leaves. In the experiments described here, plant organs were placed on the bacteria-containing solid media, and the levels of amino acids were assessed by the luminescence response of the auxotrophic bacteria. The incubation period of the plant organ/tissues with the luminescence bacteria was 1 h, followed by 5 min exposure to X-ray film. However, these two parameters may be calibrated and adjusted in cases where higher or lower detection sensitivity is required. Figure 1A displays in vivo luminescence emitted by recombinant bacteria whose viability is proportional to the free tryptophan exogenously added. A good correlation was found within the range of 20–150 µg ml–1 tryptophan. To analyse the endogenous levels of amino acids in plant tissues, specific pretreatments were tested to allow the amino acids to diffuse out and to be accessible to the bacteria. Preliminary experiments showed that storing the plant organs at –80 °C for several hours (1–12 h) followed by thawing and 1–2 h incubations on the solid bacterial medium were the range of optimal conditions for free amino acids to diffuse out of the different tissues and to reach steady-state levels in the surrounding bacterial media. However, it should be emphasized that the required step of tissue freezing may cause loss of compartmentation and may result in inaccurate determination of the subcellular localization of amino acids. Conditions may vary in different plant species or tissues and calibrating these conditions is recommended for each case. Tryptophan-requiring bacteria were chosen for this study, but similar results were obtained when two other bacterial strains requiring different amino acids were used for analysing free amino acids by the luminescence test (data not shown). This method is based on detection of a representative amino acid and is especially suitable in cases where the chosen amino acid indeed reflects the behaviour of total free amino acids.
|
|
The senescing leaf and flower were chosen as the experimental system to evaluate the suitability of this methodology for in planta analysis of free amino acids. Temporal patterns of free amino acid levels in attached and detached leaves in different monocarpic species were compared and assessed for the possible regulatory role of amino acids during leaf senescence. This study has been conducted in two different monocarpic species, oats and Arabidopsis.
Chlorophyll degradation and amino acid accumulation in detached leaves are considered to be the most prominent processes involved during senescence. Figure 2 describes the gradual decrease in chlorophyll levels which was accompanied by a parallel increase in the endogenous levels of total free amino acids during senescence of detached oat leaves (Fig. 2A, B). Similar temporal patterns of the amino acid accumulation in detached oat leaves during senescence have been found in both analysis methods: the in vivo luminescence and the colorimetric method based on the ninhydrin assay (Figs 1, 2). These results obviously reflect the characteristic intensive proteolysis occurring during senescence. The last senescence stage, 4 d incubation in darkness (Figs 1B, 2B), is, however, characterized by a reduced bioluminescence and a similar drop in total amino acids. This decrease in the levels of endogenous amino acids in the leaf is probably due to membrane leakage occurring during a late stage in senescence. Analysis of total free amino acids in the incubation medium of the detached leaves indeed revealed a significant increase in the levels of leaking amino acids during this last stage in senescence (Fig. 2C).
The temporal changes of amino acids during the progression of flower senescence were also determined by the bacterial luminescence assay. The larger tobacco flowers, which showed obvious morphological senescence changes, were chosen as a preferred and better study system compared with the tiny Arabidopsis flowers. Levels of free tryptophan progressively increased during flower senescence (Fig. 1C). Since changes in spatial distribution of amino acids within the different parts of the flowers may also occur, the upper and lower parts of the petals were divided prior to analysis and examined separately for their tryptophan content by the luminescence test (Fig. 1C). Both the upper and lower parts of the senescing tobacco flowers consistently showed an increase in the luminescence response, reflecting an increase in the endogenous tryptophan levels. This temporal profile of tryptophan changes representing free amino acids levels correlates well with the characteristic senescence-enhanced proteolysis occurring in flowers (Rubinstein, 2000).
Amino acid profiles of attached and detached leaves during senescence
Natural senescence of attached oat leaves and their amino acid content were monitored and compared with detached leaf senescence. The bacterial luminescence assay was employed to monitor the in planta temporal changes in the endogenous levels of free tryptophan in attached leaves during senescence. A constant decline in tryptophan content was found, starting from a fully developed green leaf 10 d after germination (100% chlorophyll) through the advancing stages of senescence at days 18, 32 (80, 55% chlorophyll) up to the very last phase at 42 d (20% chlorophyll) (Fig. 3A). A similar pattern of amino acid changes during development and senescence in tobacco leaves has been reported (Masclaux et al., 2000). The drop in free amino acid levels represents the dominating processes of remobilization of breakdown products from the source cells of the senescing leaf into the sink cells of growing vegetative and/or reproductive organs or tissues. The characteristic drop in photosynthetic activity (Gepstein, 1988), on the one hand, and the intensification of amino acid remobilization on the other, can explain the net decrease in the steady-state levels of free amino acids during natural leaf senescence (Fig. 2A). Rubisco, the key enzyme of the photosynthetic carbon fixation cycle, accounts for up to 35% of total nitrogen in mature leaves of C3 plants and makes a major contribution to the amino acid levels during its degradation. Rubisco levels in the leaf increased during leaf expansion and reached their maximum levels at around the time of full expansion. Thereafter, throughout the senescence stages, Rubisco levels continuously decreased and reached undetectable levels in the fully senescent leaf (Mae et al., 1983; Makino et al., 1984; Crafts-Brandner et al., 1996). The decline in Rubisco levels in senescing leaves can be attributed to a decreased rate of synthesis and/or to an increased rate of degradation. Degradation of Rubisco started at the time of the completion of leaf expansion and was most intensive during the early stages of senescence (Mae et al., 1983). During the whole senescence period of rice leaves, the degradation rate of Rubisco exceeded the rate of its synthesis and the changes in the influx and efflux of nitrogen in rice leaves was reported to correlate with the rates of synthesis and degradation, respectively, of Rubisco (Makino et al., 1984). Though Rubisco degradation contributes most to the efflux of amino acids to developing organs, the degradation of other chloroplast proteins is known to contribute to the gradual loss of photosynthetic activity during senescence (Ben-David et al., 1983; Gepstein, 1988; Bate et al., 1991). Proteins other than Rubisco thus also contribute to the free amino acid pools. Levels of the light-harvesting chlorophyll a/b binding protein (LHCPII), the most abundant protein in thylakoid membranes, stay high, however, during senescence. The reason for the high level is the suppression of LHCPII degradation rather than an increase in its synthesis (Bate et al., 1991). Other studies measuring protein synthesis and degradation by pulse labelling techniques strongly suggest that protein degradation is the major process responsible for the dramatic increase in the remobilization of nitrogen during leaf senescence. The reutilization of the remobilized nitrogen requires additional processes, such as inter-conversion of free amino acids before being transported via the phloem (Hayashi and Chino, 1990), transport from source to sink cells, and finally the conversion of the transported forms to available forms and their utilization in the sink tissues. In addition to the significant contribution of proteolysis and amino acid release in nitrogen redistribution throughout the plant during senescence, amino acid accumulation contributes to the source strength in senescing leaves, which in turn also influences considerably the rate of source to sink transport.
|
By contrast to the attached leaf that displayed a gradual decrease during the progression of the senescence, the detached oat leaf exhibited an increase in amino acid content during the progression of yellowing as defined by its chlorophyll degradation (Figs 1, 2A, B, 3B). This increase reflects the accumulation of breakdown products resulting from intense activities of various protein degradation mechanisms operating during senescence (Yoshida, 2003
). Furthermore, the sink-deprived detached leaves accumulated amino acids due to the lack of physical connection to other plant organs. The characteristic leakiness of membranes at the final stages of senescence is responsible for the decrease in the levels of endogenous free amino acid levels at the final senescence stages (Figs 1B, 2B, C, 3B).
The senescence programme of Arabidopsis, has been the focus of several recent intensive biochemical and genomics studies (Gepstein et al., 2003; Guo et al., 2004). The temporal pattern of tryptophan levels during senescence of Arabidopsis leaves was therefore also tested by the luminescence assay (Fig. 4). Like the detached oat leaf, the Arabidopsis detached leaf also showed an increase in the accumulation of free amino acids during the progression of senescence (Fig. 4B). However, by contrast to the behaviour of the oat leaf (Fig. 3A), the attached Arabidopsis leaf accumulated tryptophan throughout the senescence period (Fig. 4A). This uncommon pattern of amino acid accumulation in the attached leaf of Arabidopsis is most likely to be the result of intensive proteolytic activity and amino acid release, which is not balanced by an equal amino acid efflux from the senescing leaf.
|
Export of nutrients from the source (senescing leaves) depends not only on the intensity of the source (nutrient levels), but also on sink strength. Thus, a possible explanation for the amino acid accumulation in the attached Arabidopsis leaf is the weaker sink exerted by the small-sized vegetative and developing reproductive organs in Arabidopsis, compared with oats or legume species. It has been documented that weak nutrient sinks decrease resorption of nutrients from the senescing leaves, whereas strong sinks present during senescence increase their potential resorption. Strong sinks, which are usually found in species with large masses of maturing fruits or developing organs, should increase the potential resorption and vice versa; weak sinks should decrease this value (Killingbeck, 1996
). If the rate of source to sink transport is indeed reduced in Arabidopsis, failing to balance the rate of amino acid release in the source, the result would, inevitably, be a net amino acid accumulation in the senescing source cells.
Although the patterns of amino acid accumulation during senescence of attached leaves in Arabidopsis and oat represent opposite trends, both species display similar senescence progression patterns. Thus, the results presented here do not suggest a universal regulatory role for total free amino acids pools in triggering monocarpic senescence. In a similar way to oats, attached tobacco leaves were reported to lose amino acids, whereas sugar levels increased concomitant with the decline in photosynthetic activity during the period of the transition from sink to source leaves (Masclaux et al., 2000). These results support the notion that sugars are important regulatory molecules for triggering leaf senescence (Dai et al., 1999; Xiao et al., 2000), but there is no evidence for a similar role of amino acids during senescence.
Net accumulation of amino acids during senescence of attached Arabidopsis leaves suggests that amino acids are released faster by intensive proteolysis than they are removed by efflux from the leaf. However, this does not necessarily imply that the steady-state levels of other metabolites or factors in Arabidopsis leaves follow the same pattern during natural senescence.
|
Acknowledgements |
|---|
The critical reading of the manuscript by Dr B Horwitz is greatly appreciated.
|
References |
|---|
|
TopAbstractIntroductionMaterials and methodsResults and discussionReferences |
|---|
Adar YY, Simaan M, Ulitzur S. 1992. Formation of the luxR protein in the Vibrio fischeri-ux system is controlled by HtpR through the groESL proteins. Journal of Bacteriology 174, 7138–7143.
Bate NJ, Rothestein SJ, Thompson JE. 1991. Expression of nuclear and chloroplast photosynthesis-specific genes during leaf senescence. Journal of Experimental Botany 42, 801–811.
Ben-David H, Nelson N, Gepstein S. 1983. Differential changes in the amount of protein complexes in the chloroplast membrane during senescence of oat and bean leaves. Plant Physiology 73, 507–510.
Bhalerao R, Keskitalo J, Sterky F, et al. 2003. Gene expression in autumn leaves. Plant Physiology 131, 430–442.
Buchanan-Wollaston V, Earl S, Harrison E, Mathas E, Navabpour S, Page T, Pink D. 2003. The molecular analysis of leaf senescence – a genomics approach. Plant Biotechnology Journal 1, 3–22.
Crafts-Brandner SJ, Klein RR, Klein P, Holzer R, Feller U. 1996. Coordination of protein and mRNA abundances of stromal enzymes and mRNA abundances of the Clp protease subunits during senescence of Phaseolus vulgaris (L.) leaves. Planta 200, 312–318.[ISI][Medline]
Dai N, Schaffer A, Petreikov M, Shahak Y, Giller Y, Ratner K, Levine A, Granot D. 1999. Overexpression of Arabidopsis hexokinase in tomato plants inhibits growth, reduces photosynthesis, and induces rapid senescence. The Plant Cell 11, 1253–1266.
Dangl J, Dietrich R, Thomas H. 2000. Senescence and programmed cell death. In: Buchanan B, Gruissem W, Jones R, eds. Biochemistry and molecular biology of plants. Rockville, MD: American Society of Plant Physiologists, 1044–1100.
Feller U, Fisher A. 1994. Nitrogen metabolism in senescing leaves. Critical Reviews in Plant Science 13, 241–273.
Gepstein S. 1988. Photosynthesis. In: Nooden L, Leopold AC, eds. Senescence and aging in plants. San Diego: Academic Press, 85–109.
Gepstein S. 2004. Leaf senescence – not just a ‘wear and tear’ phenomenon. Genome Biology 5, 212.[CrossRef][Medline]
Gepstein S, Sabehi G, Carp MJ, Falah M, Hajouj T, Nesher O, Yariv I, Dor C, Bassani M. 2003. Large-scale identification of leaf senescence-associated genes. The Plant Journal 36, 629–642.[CrossRef][ISI][Medline]
Gepstein S, Thimann KV. 1981. The role of ethylene in the senescence of oat leaves. Plant Physiology 68, 349–354.
Guo Y, Cai Z, Gan S. 2004. Transcriptome of Arabidopsis leaf senescence. Plant, Cell and Environment 27, 521–549.[CrossRef]
Hayashi H, Chino M. 1990. Chemical-composition of phloem sap from the uppermost internode of the rice plant. Plant and Cell Physiology 31, 247–251.
He Y, Gan S. 2002. A gene encoding an acyl hydrolase is involved in leaf senescence in Arabidopsis. The Plant Cell 14, 805–815.
Hensel LL, Nelson MA, Richmond TA, Bleecker AB. 1994. The fate of inflorescence meristems is controlled by developing fruits in Arabidopsis. Plant Physiology 106, 863–876.[Abstract]
Himelblau E, Amasino RM. 2001. Nutrients mobilized from leaves of Arabidopsis thaliana during leaf senescence. Journal of Plant Physiology 158, 1317–1323.[CrossRef][ISI]
Hortensteiner S, Feller U. 2002. Nitrogen metabolism and remobilization during senescence. Journal of Experimental Botany 53, 927–937.
Killingbeck KT. 1996. Nutrients in senesced leaves: keys to the search for potential resorption and resorption proficiency. Ecology 77, 1716–1727.[CrossRef]
Lers A, Khalchitski A, Lomaniec E, Burd S, Green PJ. 1998. Senescence-induced RNases in tomato. Plant Molecular Biology 36, 439–449.[CrossRef][ISI][Medline]
Mae T, Makino A, Ohira K. 1983. Changes in the amounts of ribulose bisphosphate carboxylase synthesized and degraded during the life-span of rice leaf (Oryza sativa L.). Plant and Cell Physiology 24, 1079–1086.
Makino A, Mae T, Ohira K. 1984. Relation between nitrogen and ribulose-1,5-bisphosphate carboxylase in rice leaves from emergence through senescence. Plant and Cell Physiology 25, 429–437.
Masclaux C, Valadier MH, Brugiere N, Morot-Gaudry JF, Hirel B. 2000. Characterization of the sink/source transition in tobacco (Nicotiana tabacum L.) shoots in relation to nitrogen management and leaf senescence. Planta 211, 510–518.[CrossRef][ISI][Medline]
Nam HG. 1997. The molecular genetic analysis of leaf senescence. Current Opinion in Biotechnology 8, 200–207.[CrossRef][ISI][Medline]
Nooden L. 1980. Senescence in the whole plant. In: Thimann K, ed. Senescence in plants. Boca Raton, FL: CRC Press, 219–258.
Nooden LD, Penney JP. 2001. Correlative controls of senescence and plant death in Arabidopsis thaliana (Brassicaceae). Journal of Experimental Botany 52, 2151–2159.
Okada K, Katoh S. 1998. Two long-term effects of light that control the stability of proteins related to photosynthesis during senescence of rice leaves. Plant and Cell Physiology 39, 394–404.
Ono K, Ishimaru K, Aoki N, Ohsugi R. 1999. Transgenic rice with low sucrose-phosphate synthase activities retain more soluble protein and chlorophyll during flag leaf senescence. Plant Physiology and Biochemistry 37, 949–953.[CrossRef]
Quirino BF, Normanly J, Amasino RM. 1999. Diverse range of gene activity during Arabidopsis thaliana leaf senescence includes pathogen-independent induction of defense-related genes. Plant Molecular Biology 40, 267–278.[CrossRef][ISI][Medline]
Rubinstein B. 2000. Regulation of cell death in flower petals. Plant Molecular Biology 44, 303–318.[CrossRef][ISI][Medline]
Terce-Laforgue T, Mack G, Hirel B. 2004. New insights towards the function of glutamate dehydrogenase revealed during source–sink transition of tobacco (Nicotiana tabacum) plants grown under different nitrogen regimes. Physiologia Plantarum 120, 220–228.[CrossRef][Medline]
Thompson JE, Froese CD, Madey E, Smith MD, Hong Y. 1998. Lipid metabolism during plant senescence. Progress in Lipid Research 37, 119–141.[CrossRef][ISI][Medline]
Tozawa T, Hasegawa H, Terakawa T, Wakasa K. 2001. Characterization of rice anthranilate synthase 
-subunit genes OASA1 and OASA2: tryptophan accumulation in transgenic rice expressing a feedback-insensitive mutant of OASA1. Plant Physiology 126, 1493–1506.
Ulitzur S. 1997. Established technologies and new approaches in applying luminous bacteria for analytical purposes. Journal of Bioluminescence and Chemiluminescence 12, 179–192.[CrossRef][ISI][Medline]
van Doorn WG. 2004. Is petal senescence due to sugar starvation? Plant Physiology 134, 35–42.
Xiao W, Sheen J, Jang JC. 2000. The role of hexokinase in plant sugar signal transduction and growth and development. Plant Molecular Biology 44, 451–461.[CrossRef][ISI][Medline]
Yoshida K. 2003. Molecular regulation of leaf senescence. Current Opinion in Plant Biology 6, 79–84.[CrossRef][ISI][Medline]
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  E. van der Graaff, R. Schwacke, A. Schneider, M. Desimone, U.-I. Flugge, and R. Kunze Transcription Analysis of Arabidopsis Membrane Transporters and Hormone Pathways during Developmental and Induced Leaf Senescence Plant Physiology, June 1, 2006; 141(2): 776 - 792. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Veljovic-Jovanovic, B. Kukavica, B. Stevanovic, and F. Navari-Izzo Senescence- and drought-related changes in peroxidase and superoxide dismutase isoforms in leaves of Ramonda serbica J. Exp. Bot., May 1, 2006; 57(8): 1759 - 1768. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||