JXB Advance Access originally published online on January 31, 2006
Journal of Experimental Botany 2006 57(4):801-814; doi:10.1093/jxb/erj063
RESEARCH PAPER |
Diel patterns of leaf C export and of main shoot growth for Flaveria linearis with altered leaf sucrose–starch partitioning
Department of Plant Agriculture (Bovey Building), University of Guelph, 50 Stone Road E, Guelph, Ontario, N1G 2W1 Canada
* To whom correspondence should be addressed. E-mail: bgrodzin{at}uoguelph.ca
Received 14 July 2005; Accepted 15 November 2005
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Diel C export from source leaves of two Flaveria linearis lines [85-1: high cytosolic fructose-1,6-bisphosphatase (cytFBPase) and 84-9: low cytFBPase] were estimated using three methods, including leaf steady-state 14CO2 labelling, leaf metabolite analysis, and leaf dry mass analysis in conjunction with leaf CO2 exchange measurements. Synthesis and accumulation of starch during the daytime were much higher in 84-9. Relative 14C-export (export as a % of photosynthesis) in the light was 36% higher in 85-1. The diel export patterns from 14C-analyses correlated with those based on metabolite or dry weight/gas exchange analyses during the daytime, but not during the night. Night-time export estimated from 14C-disappearance was 3.6 times lower than those estimated using the other methods. Even though the starch degradation at night was greater for 84-9, night-time export in 84-9 was similar to 85-1, since 84-9 showed both higher respiration and accumulation of soluble sugars (i.e. glucose) at night. Patterns of 14C allocation to sink organs were also different in the two lines. Main stem growth was less in 84-9, being reduced most in the light when leaf export was lower relative to 85-1. Supplementation with sucrose for 1 h daily via the roots at a time when leaf export in 84-9 was low relative to 85-1 increased the stem growth rate of 84-9 to a level similar with that of 85-1. This study provides evidence that diel C availability predicted by source strength (e.g. C-export rate) influences main stem extension growth and the pattern of sink development in F. linearis.
Key words: C export, C partitioning, C3–C4 intermediates, cytosolic fructose-1,6-bisphosphatase, diel patterns, Flaveria, photosynthesis, shoot growth, sucrose, starch
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
In addition to being a source of energy and C, recent studies have shown that sugars are also involved in regulating developmental responses such as flowering and vegetative development through mechanisms that sense and transduce sugar status (Sheen et al., 1999
; Moore et al., 2003). However, studies demonstrating sugar sensing and transduction of a sugar signal into a physiological response in planta are lacking and few studies have examined the potential involvement of diel (i.e. day and night) sugar fluxes (e.g. sucrose export rates) in sugar sensing (Kehr et al., 1998). The importance of understanding the role that different day and night export patterns may have on growth and development is illustrated by studies with genetic variants that show the plasticity of leaf starch and sugar metabolism (Caspar et al., 1985, 1991; Lin et al., 1988; Frommer and Sonnewald, 1995; Stitt and Sonnewald, 1995; Zrenner et al., 1996; Hausler et al., 1998; Kehr et al., 1998; Zeeman et al., 1998; Zeeman and ap Rees, 1999; Strand et al., 2000; Nielsen and Stitt, 2001). Although development and growth in some genetic variants is not affected (Zrenner et al., 1996; Hausler et al., 1998; Nielsen and Stitt, 2001), in other cases growth is inhibited (Riesmeier et al., 1993; Strand et al., 2000) and development of these genetic variants is similar to that of the wild-type controls only in continuous light (Caspar et al., 1991; Zeeman et al., 1998). In addition, a number of developmental processes including vegetative and floral stem elongation, leaf expansion, and fruit expansion show diel or circadian patterns of growth (Kristie and Jolliffe, 1985; Fishman and Genard, 1998; Jouve et al., 1998; Schmundt et al., 1998). It can be hypothesized that the diel availability of recently fixed C may impact on developmental processes, and particularly those processes that vary in expression in a diel or circadian manner.
Different CO2 concentrating and fixation mechanisms have evolved (e.g. C3, C3–C4 intermediate, and C4 types) that are reliant on complex intercellular movements of metabolites in the leaf (Hatch, 1987; Edwards and Ku, 1987). The type II C3–C4 intermediate species (e.g. F. linearis) have relatively high leaf photosynthesis rates due to reduced photorespiratory loss of CO2 and are thought to represent an evolutionary step from C3 to C4 metabolism (Monson et al., 1986; Edwards and Ku, 1987; Ku et al., 1991; Dai et al., 1996). In a earlier study, F. linearis source leaves exported less C during photosynthesis than did leaves of other Flaveria C3 and C4 species and accumulated more starch than sugars at the end of the light period (Leonardos and Grodzinski, 2000). In addition, F. linearis leaves actually mobilized a large amount of carbon from sugar and starch reserve pools at night, but did not fully compensate for their slow export during photosynthesis (Leonardos and Grodzinski, 2003).
Integrated control of C metabolism leads to balanced synthesis of starch in the chloroplasts and sugars (mainly sucrose) in the cytosol. During the daytime, starch accumulates when sucrose synthesis exceeds the export capacity of the leaf. Decreased flux of C through sucrose synthesis may lead to lower export and higher accumulation of starch during the daytime. This starch is utilized during the night to sustain respiration and export. Alterations in initial C partitioning in the leaf may affect the availability of translocates (i.e. sucrose), day/night export patterns, and even growth and development (Micallef et al., 1995; Kehr et al., 1998; Grodzinski et al., 1999). Plants differing in leaf sucrose–starch partitioning provide a potential means to study the effects of altered diel sucrose export rates. For example, a naturally occurring mutant of F. linearis has been found with reduced activity of cytosolic fructose-1,6-bisphosphatase (cytFBPase) and altered C partitioning (Sharkey et al., 1992; Micallef and Sharkey, 1996). Cytosolic fructose-1,6-bisphosphotase (cytFBPase) is a key enzyme in the C metabolism pathway regulating the flow of C from triose-P into the hexose-P pools and the synthesis of sucrose (Stitt et al., 1987; Daie, 1993). Photosynthesis and C partitioning have been studied in these F. linearis plants (Sharkey et al., 1992; Micallef and Sharkey, 1996) as well as in transgenic plants with antisense repression of cytFBPase (Zrenner et al., 1996; Strand et al., 2000, 2003). Sharkey et al. (1992) hypothesized that the mutant plants survive the reduced cytFBPase activity by partitioning more C to starch than to sucrose during the day and remobilizing the excess starch at night. The same was concluded for transgenic Arabidopsis plants by measuring changes of carbohydrate pools during a day and night cycle (Strand et al., 2000). However, there are no studies that have accurately quantified export flux rates of newly fixed C from source leaves with reduced cytFBPase activity during a day and night cycle.
Various methods have been used to quantify diel C export rates from leaves, including: (i) diel leaf dry weight/C quantification coupled with diel net CO2 exchange rates (Method 1) (Goodall, 1946; Terry and Mortimer, 1972; Ho, 1978); (ii) diel leaf metabolite quantification coupled with diel net CO2 exchange rates (Method 2) (Galtier et al., 1995); and (iii) use of continuous 14CO2 labelling coupled with either quantification of 14C export by multiple sampling or by using a Geiger Müller (GM) detector (Method 3) (Geiger and Fondy, 1979; Hendrix and Grange, 1991; Grodzinski et al., 1998). For Method 3 to be accurate, isotopic equilibrium (i.e. steady-state conditions) has to be reached for all relevant metabolite pools, and isotopic discrimination cannot be occurring for relevant reactions. Isotopic equilibrium may not be reached in 14C labelling if pools other than those involving recently fixed C are contributing to C for export. However, Method 3 does provide information on the export of recently fixed C, and it allows for continuous monitoring of C export rates non-destructively. Method 2 requires that all relevant metabolites/end products have been assayed and it involves destructive harvesting; it is usually assumed that starch, neutral sugars, and CO2 constitute the major metabolites. Method 1 has the same requirements as Method 2, but Method 1 by its nature is more inclusive in the total metabolites assayed; however, Method 2 provides more specific information on the partitioning of C into different pathways and metabolites. No single study to date has combined and compared all of these methods for quantifying diel C export patterns in a species.
In this study, export fluxes of C during a day and night cycle from F. linearis leaves differing in sucrose–starch partitioning have been quantified using the three methods described above, and correlations with diel growth patterns of the whole plant were examined, including stem elongation which is known to show a circadian growth pattern. For the 14C method (i.e. Method 3), the rates of 14C-export from mature attached leaves were measured during a steady-state feed period in the light and during a chase period in the dark. 14C-partitioning within the fed leaf at the beginning (i.e. end of day) and at the end (i.e. end of night) of the chase period were determined to assess the extent of breakdown of the major pools accumulated during the daytime feed. 14C-allocation among the different plant parts was also examined to determine the diel C sink demands within the whole plant. Estimates of diel C export rates using Method 2 were also obtained from two independent experiments as follows: (i) by quantifying diel pool sizes of neutral sugars and starch, and determining diel net CO2 exchange rates at two time points in the 14C experiment; and (ii) by quantifying diel pool sizes for neutral sugars and starch at ten time points over 24 h coupled with the determination of diel net CO2 exchange rates on a different set of plants. C export was also quantified by measuring diel changes in leaf dry weights at three time points coupled with net CO2 exchange rates in a separate experiment (Method 1). These data were compared against diel patterns of main stem elongation rates and growth. Finally, the effect of a 1 h sucrose application through the roots starting at noon on main stem extension growth was examined.
|
Materials and methods |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Plant material and growing conditions
F. linearis lines 85-1 (wild type; high activity of cytFBPase) and 84-9 (mutant; low activity of cytFBPase) were grown from rooted cuttings in growth chambers at 200 µmol m–2 s–1 PPFD, ambient CO2 (37 Pa), 16/8 h (day/night) photoperiod, and 25 °C (day/night) in either soilless mix or in a sterile hydroponic system as indicated in the figure legends.
Leaf dry weight measurements (Method 1)
To determine diel leaf dry weights, 0.5 g of fresh leaf tissue was sampled per plant from a total of 16 plants per line in two separate experiments at three time points (beginning of light period, end of light period, end of dark period) and dry weights were determined. Some leaf tissue at each time point was also sampled and the leaf areas determined by tracing the leaves on transparency film, and then cutting out the traces to determine the area relative to a standard. This leaf tissue was dried in a forced air oven at 60 °C until no further changes in dry weight were measured. A dry weight/leaf area ratio was determined for each treatment.
Leaf C metabolite and enzyme activity measurements (Method 2)
The determination of diel leaf metabolite levels (starch and sugars) were performed for a set of plants sampled 10 times over a 24 h period. For this multiple sampling experiment, starch and sugars in leaves were quantified using enzyme linked assays as described by Sharkey et al. (1992).
The diel activity of cytFBPase was quantified as described by Micallef and Sharkey (1996) with the following modification. The rate of FBP hydrolysis was determined with and without Mg2+ in the reaction mixture, and cytFBPase activity was calculated as follows: cytFBPase activity=activity with Mg2+–activity without Mg2+. Experiments using polyclonal antibodies raised to carrot root aldolase (provided by Bill Plaxton, Queen's University, Kingston, Ontario, Canada) have confirmed that Mg2+-independent FBP hydrolysis activity in Flaveria linearis is catalysed by aldolase (BJ Micallef, unpublished data). The assay of pyrophosphate-dependent phosphofructokinase activity (PPi-PFK) was as described by Yaun et al. (1988), and assayed in the direction of FBP formation using 5 mM F6P, 2.5 mM PPi, 5 mM MgCl2, and 1 µM F26P2; tissue was sampled in the middle of the light period.
Leaf gas exchange measurements (Methods 1 and 2)
A Li-Cor 6400 portable gas exchange unit (Li-Cor, Lincoln, Nebraska, USA) was used for determinations of diel net CO2 exchange rates in these experiments. Plants were measured in the growth chamber used for growth using only the lighting from the chamber at a leaf temperature of 25 °C and an average relative humidity of approximately 60% (VPD=1.1 kPa). Unshaded and fully expanded leaves near the top of the plant were chosen. Daytime and night-time rates were determined by randomly sampling one leaf per plant over the day or night period, respectively, and measuring each leaf for a period of 10–15 min or until a stable rate was obtained. The results over the day or night were averaged to obtain the net CO2 exchange rates.
Leaf 14C-export and gas exchange measurements (Method 3)
14CO2 steady-state labelling and gas exchange analysis on the first–second attached fully expanded leaf from the apex (i.e. 1–2 internodes and nodes above the fed leaf) on 18–25-d-old plants were conducted with an open-flow system during a 16-h feed in the light and an 8 h chase in the dark as described previously by Leonardos et al. (2003). Immediate export rates during the daytime were determined starting 60–90 min after the feed began, while isotopic equilibrium existed between the 14CO2 available for photosynthesis and the 14C-translocates (Leonardos and Grodzinski, 2000, 2001). The level of 14C in the leaf was monitored with a GM detector during the feed and chase period correcting for the efficiency of the GM detector. The total 14C-export during the daytime was estimated as the difference between the total 14C assimilated and that remaining in the leaf at the end of the feed; the total daytime C export rates also correspond to the area under the curves in Fig. 2B. 14C-export during the night-time was corrected for respiratory release of 14CO2 in the dark. Dark 14C-respiration was determined by trapping the outlet gas in 100 ml of ethanolamine/ethylene glycol monomethyl ether (1:2 v:v) and counting the radioactivity in the trapping solution by liquid scintillation counting (Leonardos et al., 1996).
|
Leaf 14C-partitioning and whole-plant 14C-allocation patterns
To identify those pools of labelled intermediates which were mobilized during the chase periods, 14C-partitioning in source leaves was determined at the end of the 16 h feed and of the entire 24 h feed and chase experiment as described previously (Leonardos and Grodzinski, 2000
). One set of plants was harvested at the end of the daytime feed (23.00 h) and another at the end of the night-time chase (07.00 h) to determine diel 14C-allocation patterns within the plant. Each plant was divided into sections according to their position in relation to the fed leaf (i.e. leaves above, stem above, laterals above, leaves below, stems below, laterals below, and roots). Plant parts were oven-dried for 48 h and 14C-content was measured by liquid scintillation after combustion of the samples using a biological oxidizer (BIO-OX, RJ Harvey Instrument Co., Hillsdale, NJ, USA).
Main stem extension growth
Diel main stem extension rates and patterns were determined non-destructively using a rotary motion sensor (model CI-6625, Pasco Scientific, Roseville, CA, USA) which has a resolution of 20 µm. The motion sensor was connected to a Vernier LabPro Datalogger (Vernier Software and Technology, Beaverton, OR, USA). Using a Texas Instruments Model TI-83 plus calculator (Texas Instruments Inc., Dallas, Texas, USA), data were downloaded using TI-Graph LinkTM for Windows TI-83 plus version V2.3. The data were then uploaded into IGOR Pro Version 3.14 from WaveMetrics, Inc. (Lake Oswego, Oregon, USA) to allow graphing of data and determinations of stem extension rates and total stem extension. Measurements on any plant lasted up to 12 d as it takes this long for an internode to extend fully in F. linearis; stem extension typically remained linear over at least 8 d.
The rate of main stem internode elongation for lines 84-9 and 85-1 in mg C d–1 was estimated by sampling the top main stem internode from a set of six plants per line (30 plants per line in total) every 2 d over a period of 8 d, which were then dried down. Internode lengths and widths were also measured at each time point. The average length of the internodes on all plants at time 0 was 1 mm.
Sucrose supplementation through roots
A sterile apparatus was developed to allow supplementation of sucrose via the roots for a period of 1 h daily. The apparatus consisted of a 135 ml glass jar (5.5 cm in diameter and 7.5 cm high to the rim) covered with a Magenta top (Magenta Corp, Chicago, USA) that was fitted with a 1.5 cm wide silicone stopper (Fisherbrand, No. 30, Cat # FB57896) cut out on the bottom to accommodate the plant stem, and a 9 mm IceBlueTM septa (Cat # 22381, Restek Corporation, 814-353-1300) for inserting a syringe needle. Plant cuttings were made and inserted through the silicone stopper so that the stem was in 100 ml of one-quarter strength Hoagland's solution, cotton was placed between the stem and the depression in the silicone stopper, and plants were covered with plastic wrap to prevent wilting. During rooting, fresh solution was added every 2 d. Once the plants were sufficiently rooted (around 10 d later), the apparatus was transferred to a sterile hood and the Hoagland's solution removed, and the roots and other exposed surfaces were treated for 1 min using a 1/100 dilution of 5.25% sodium hypochlorite bleach. Preliminary experiments using plates containing either Plate Count Agar (PCA) or PCA plus 25 µg ml–1 chloramphenicol (PCA-C) showed that this treatment killed all plateable bacteria and fungi present in the rooting solution. After the 1 min treatment, the bleach solution was removed, all surfaces were rinsed with sterile deionized water, and a piece of sterile cotton was placed in the silicone stopper depression. The cotton was used both to act a microbial filter and the cotton fibres helped to keep the plant fixed in place. At this point, the Magenta top was never removed from the glass jar until the termination of the experiment. One hundred and five ml of sterile half-strength Hoagland's solution was then pumped into each jar. This was done using a sterilized (i.e. surfaces cleaned with bleach and rinsed with sterile water) medium-to-high flow Fisherbrand peristaltic pump (Model 3389, flow rate 4.0–600 ml min–1) fitted with a 7.5 cm long Hamilton syringe needle; the needle was dipped in dilute bleach and then inserted into the IceBlueTM septa. Plants were then moved to the growth chamber. Due to the bleach treatment, the plants remained slightly wilted for 1 d but recovered fully within 2 d. At this point, the sugar supplementation treatments were started.
At 11.00 h daily, the root media was injected with a sterile 0.5 M sucrose solution to a final concentration of 2.5 mM using a sterile plastic syringe fitted with a 16 gauge needle. Each plant typically used 3–5 ml water d–1, bringing the initial volume close to 100 ml. After 1 h, the 2.5 mM sucrose solution was pumped out using the peristaltic pump and 105 ml of fresh sterile half-strength Hoagland's solution was pumped into the jar. The plants were treated with sucrose over a 7 d period, and then the rotary motion sensors were attached to the terminal growing point using a piece of thread that was then draped around the pulley of the motion sensor and the other end weighted using a small metal washer. Sucrose treatments then continued for 8–9 d.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Leaf enzyme activities and C metabolites and fluxes (Methods 1 and 2)
For the Method 2 experiment, the activity of cytFBPase in leaves remained constant throughout the day and averaged 1.70±0.20 and 0.02±0.01 µmol FBP hydrolysed m–2 s–1 for lines 85-1 and 84-9, respectively (Fig. 1A). The PPi-PFK activities for lines 84-9 and 85-1 in the middle of the light period were 3.5±0.1 and 2.1±0.2 µmol FBP m–2 s–1, respectively. Similar levels of sucrose were found in both lines during either the day or the night period (Fig. 1B), but both glucose and fructose levels increased starting near the beginning of the dark period for line 84-9 (Fig. 1C, D). Starch synthesis rates during the day were higher in 84-9 (3.7±0.3 versus 1.8±0.2 µmol C m–2 s–1 for 85-1; calculated from Fig. 1E), and as a result, starch levels in 84-9 were much higher at the end of the daytime (Fig. 1E). More starch was metabolized during the night in 84-9, but still more starch remained in 84-9 at the end of the night (Fig. 1E). The photosynthetic rate for line 84-9 was 90% of the rate for line 85-1, although line 84-9 had a calculated daytime C export rate that was only 50% of line 85-1 (Table 1). The calculated sucrose synthesis rates during the day for 84-9 and 85-1 were 1.8±0.2 and 4.3±0.3 µmol C m–2 s–1, respectively (calculated as daytime net C assimilation rate–starch synthesis rate).
|
|
In the Method 1 experiment, the photosynthetic rates for lines 84-9 and 85-1 were 5.9±0.4 and 5.1±0.3 µmol C m–2 s–1, and the net changes in dry weight between the end of day and end of night were 3.9±0.4 and 3.1±0.3 g, respectively. The calculated daytime C export rates for lines 84-9 and 85-1 in the diel dry weight experiment were 4.1±0.8 and 2.7±0.3 µmol C m–2 s–1, respectively.
Leaf 14C-export and net CO2 exchange (Method 3)
Diel patterns of net C exchange rate (NCER) and 14C export in source leaves were determined in lines 84-9 and 85-1 at similar stages of development (Fig. 2). At the growth conditions (i.e. 200 µmol m–2 s–1 PPFD, 37 Pa CO2, and 25 °C), photosynthesis and export rates during the daytime were higher in 85-1 than in 84-9 (Fig. 2A, B). In both lines, leaf photosynthesis peaked after 2–3 h in the light (Fig. 2A) and remained relatively constant until the end of the daytime period. Photosynthesis rates in the wild type decreased gradually towards the end of the light period to levels similar to those of the mutant. The average photosynthetic rates for lines 84-9 and 85-1 were 4.9 and 5.6 µmol C m–2 s–1, respectively, and the respiratory rates were 0.9 and 0.8 µmol C m–2 s–1, respectively (Fig. 2A). Export rates in both lines appeared to level off after 3–4 h in the light, but started to increase again during the last 3–4 h of the photoperiod (Fig. 2B). As absolute values, export rates in the light from the wild type was about 40% higher than that from the mutant leaves (Fig. 2B). Export increased relative to photosynthesis in both lines throughout the daytime (Fig. 2C). In the wild type leaves, 60–80% of the newly fixed 14C was transported immediately for most of the day. By comparison, in the mutant leaves export in the light was only 40–60% of the photosynthetic rate. Interestingly, export rates were higher at the beginning of the dark period in 84-9, but similar in the two lines at the end of the night (Fig. 2B).
Figure 3 shows the fate of the newly fixed 14C in source leaves of both lines during the 24 h day/night period. The total of all bars within each plant line equals the total amount of 14C fixed during the feed period. As shown above in Fig. 2, more 14C was exported during the light than during the dark in both lines (Fig. 3). Total export during the daytime was 40% lower in 84-9. This lower export during the daytime in the mutant was partially compensated by a 15% higher export of 14C during the night. Nevertheless, more 14C remained in the leaves of 84-9 both at the end of the 24 h period and more 14C was respired over night from 84-9. The respiration rates determined from the 14C data were about 50% of the rates determined from the net C exchange data for both plant lines (compare Figs 2A, 3). In total, more 14C was exported in the wild type over the whole day (Fig. 3).
|
Leaf C partitioning in the 14C-labelling experiment
At the end of the light period, the mutant had accumulated more than twice the amount of total starch found in leaves of the wild type (Fig. 4A). Sucrose levels in the mutant were slightly less than those in the wild type. By the end of the dark period, starch pools were significantly reduced in both lines (Fig. 4B). Sucrose levels also declined. Glucose levels were higher in 84-9 leaves than in the wild type at the end of both the light and dark periods. The total neutral sugar levels increased in line 84-9 during the dark period, but they declined for line 85-1.
|
Partitioning of newly fixed 14C (Fig. 4C, D) resembled that of the total C pools (Fig. 4A, B). In both lines, 14C was mainly in starch at the end of day (Fig. 4C). More 14C-starch was recovered in 84-9 than in 85-1 (Fig. 4C). 14C-starch supported most of the export during the night in both lines (Fig. 4C, D). Mobilization of additional starch during the night was reflected by the dark respiration rate, which was higher in 84-9 (Figs 1A, 3). 14C-sucrose also was reduced overnight. 14C-fructose levels were similar in both lines at the end of the light period and did not change during the night. 14C-glucose levels were higher in the mutant during light and further increased at the end of the dark (Fig. 4C, D). Elevated 14C-glucose levels in 84-9 may reflect increased starch degradation. In both lines, 14C-amino acids and organic acids levels were similar accounting for about 5–10% each of the total 14C remaining at the end of the day (data not shown). 14C-amino acids and organic acids decreased in both lines by the end of the night. This decrease in 14C-amino acids in 84-9 was higher than that in 85-1.
The total decrease in 14C-ethanol-insoluble metabolites between the end of day and end of night was accounted for by the loss of 14C in starch (compare Fig. 4C, D with Fig. 4E, F); thus, there was no net change in other 14C-ethanol insolubles during the dark period. The total 14C-water-soluble pool for both lines decreased similarly, and there was little net change in 14C-chloroform-soluble metabolites in the dark (Fig. 4E, F).
Comparison of diel C fluxes
Day and night-time C export rates were estimated for all experiments/methods (Table 1). The night-time export rates determined from continuous monitoring using a GM counter gave rates that were, on average, 3.8 and 3.6 times lower for lines 84-9 and 85-1, respectively, compared with the other three estimates. The other three estimates of night-time export were fairly consistent for both lines. Overall, the night-time C export rate was only slightly higher for line 84-9. The daytime C export rate averaged over experiments for line 84-9 was 64% of the rate for line 85-1; the absolute rates of daytime C export were fairly consistent among experiments, although slightly lower for the 14C estimates in nearly all cases. Averaged over experiments, daytime C export for lines 84-9 and 85-1 accounted for 56% and 70% of total C export, respectively. Photosynthetic rates, on average, were 13% lower for line 84-9 relative to line 85-1, and respiratory rates were 36% higher in line 84-9. The average daily leaf net C exchange rate for line 85-1 ((334–21) mmol C m–2=313 mmol C m–2) was 19% greater relative to line 84-9.
Whole plant growth and 14C-allocation patterns
The allocation of 14C within the plant at the end of the light and at the end of the dark is indicative of the activity of the sinks during those periods (Fig. 5). The fed source leaf supplied 14C-photoassimilates to both plant parts above and parts below its position within the plant in both lines. At the end of the light period, 14C was evenly allocated between parts above and below the fed leaf (Fig. 5A), whereas night-time allocation appeared to shift towards parts below (e.g. roots) especially in the mutant (Fig. 5D). Leaves above the fed leaf in both lines were strong sinks and acquired significant amounts of 14C. More 14C was found in the leaves above in 85-1 than in 84-9 at the end of the light period, but equal amounts at the end of dark (Fig. 5B, E). Lateral shoots above also attracted a large amount of 14C. On a per dry mass basis, they were the strongest sink (Fig. 5C, F). Lateral shoots above in the wild type had more 14C than the mutant at both the end of the light and the end of dark period (Fig. 5B, E). More 14C was metabolized in the lateral shoots above in 84-9 than in 85-1. Overall, line 85-1 accumulated significantly more 14C above the fed leaf during the daytime compared with 84-9 on both a total and dry mass basis (Fig. 5).
|
In both lines, very little 14C was found in the mature source leaves below the fed leaf (Fig. 5B, E). Also, lateral shoots below appeared to be a weak sink, which may indicate that their growth was supported by lower source leaves or they are less metabolically active (Fig. 5C, F). Lateral shoots below in 84-9 acquired more 14C than those in 85-1 in spite of their lower mass. In both lines, the majority of the 14C in the lower parts of the plant was found in the stem below and the roots (Fig. 5B, E). The roots in both lines attracted a significant amount of 14C especially during the night, however, on a per dry mass basis they were the weakest sink (Fig. 5C, F). About 30.25±5.39% and 24.96±2.65% of the 14C assimilated during the feed period was respired by the next morning in lines 85-1 and 84-9, respectively.
Main stem extension growth
Extension rates and mass gain of the main stem were higher by 56% and 100%, respectively, for line 85-1 relative to line 84-9 (Table 2). The wild type showed more than double the main stem extension growth during the first half of the photoperiod (07.00–15.00 h). During the second half of the light period (15.00–23.00 h), stem extension was at a minimum in both lines. Main stem elongation showed a typical circadian pattern for both lines, with a peak close to the end of night and a trough at around 19.00 h (data not shown). Main stem growth in 84-9 did not compensate during the night period, although approximately 40% of main stem growth for both lines occurred during this period. Internode elongation was linear over the 8 d period and widths reached a maximum of 2 mm prior to the 8 d, indicating that only primary growth occurred during this time frame.
|
The average rates of main stem extension growth for lines 84-9 and 85-1 when not supplemented with sucrose in the sterile hydroponic apparatus were 1.91±0.14 mm d–1 and 3.69±0.27 mm d–1 (calculated from the incremental growth increases as shown in Fig. 6), respectively. These are similar to the rates found for plants grown in soilless mix (Table 2). Supplementation with sucrose for 1 h daily starting at 11.00 h increased the main stem growth rate for line 84-9 close to the rate found for line 85-1 (i.e. the rate for line 84-9 increased from 1.91 to 3.31±0.19 mm d–1, or a 73% increase relative to the control). There was a 4 d lag in the response of line 85-1 to supplemental sucrose, after which the rate increased relative to the 85-1 control. The average growth rate for line 85-1 supplemented with sucrose was 4.64±0.92 mm d–1, or a 26% increase relative to the control. Supplementation with sucrose also altered the diel growth pattern of the main stem in both lines, and the effect was greatest during the period when sucrose was added (i.e. period 11.00–15.00 h) (Table 3). The effect was also greatest for line 84-9. During the daily period when sucrose was supplemented (i.e. period 11.00–15.00 h), the percentage of daily growth that occurred during this period increased 2 times for line 84-9 and 1.5 times for line 85-1. The absolute extension rate during period 11.00–15.00 h increased from 0.21 mm d–1 to 0.66 mm d–1 for line 84-9 when supplemented with sucrose, which is close to the rate of 0.58 mm d–1 during period 11.00–15.00 h found for line 85-1 without sucrose. The internode lengths for internodes 2 and 3 (those internodes that reached their final length during the course of sucrose supplementation) were 1.7±0.2 cm for both line 84-9 with or without sucrose, and 1.9±0.1 and 2.1±0.1 cm for line 85-1 with or without sucrose, respectively.
|
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Estimating leaf C export
Evidence is provided showing that altered sucrose–starch partitioning in the leaf can alter diel patterns of C export in F. linearis. The effect of increased partitioning to starch in line 84-9 was most pronounced in the daytime, when the C export rate was only 64% of that found for line 85-1 (Fig. 2; Table 1). Interestingly, although significantly more starch was degraded at night in line 84-9, the night-time C export rate was not significantly greater compared with line 85-1. There were two reasons for this observation: (i) respiration rates were up to 15 mmol C m–2 greater at night in line 84-9 (Figs 2, 3; Table 1); and (ii) in particular, accumulation of neutral sugars during the night period in line 84-9 were up to 44 mmol m–2 higher than in line 85-1 (Fig. 1). Stored reserves are used at night for maintenance respiration and also to sustain phloem loading and translocation, making export at night a more costly process than in the light (Geiger and Servaites, 1994
; Bouma et al., 1995; Noguchi et al., 2001). Since the neutral sugars are depleted during the light period in line 84-9, it is possible that some C that it not recently fixed will be exported from the leaf during the subsequent light period. In support of this hypothesis, the daytime export rate in the 14C-labelling experiment estimated from metabolite levels was 46 mmol C m–2 greater than that estimated using the GM counter (Table 1). However, the GM counter estimate only measures recently fixed C, whereas the metabolite estimate encompasses both previously and recently fixed C. Therefore, some compensation in diel C export patterns can occur in response to altered leaf sucrose–starch partitioning.
It should be noted that these measurements were done under low irradiance levels, lower than those of Micallef et al. (1996), but more importantly at sub-saturating levels for photosynthesis. This was a conscious decision to keep the absolute rates of photosynthesis as close as possible between lines, since the authors were interested in examining diel C partitioning patterns, irrespective of differences in photosynthetic rates. As environmental conditions become more favourable for maximum photosynthetic rates, the photosynthetic rate of line 85-1 (high cytFBPase) relative to line 84-9 (low cytFBPase) increases (Micallef et al., 1996). In fact, the maximal sucrose synthetic rate for line 84-9 is about 3 µmol C m–2 s–1 (BJ Micallef, unpublished data), which is close to that found in the present study. Therefore, as the photosynthetic rates increase, greater differences in sucrose-starch partitioning and diel C export patterns would be predicted between lines 84-9 and 85-1.
This is the first study to compare currently available methods for estimating diel C export patterns in plants. Overall, the methods used gave relatively consistent results for daytime C export, although the 14C-continuous monitoring method appeared slightly to underestimate the daytime rate as discussed above (Table 1). However, the night-time C export rates estimated using the 14C-continuous monitoring method were up to 3.8 times lower relative to the other methods. One possibility is that both labelled and unlabelled starch was degraded during the night period. Comparison of night-time respiration rates determined by either gas exchange or 14C-monitoring provides direct evidence for this proposal. In particular, the respiration rates for both lines as determined by gas exchange were double the rates estimated from the 14C-data (compare Figs 2, 3). The only explanation is that isotopic dilution is occurring for the C being shunted into the respiratory reactions at night. Where would this unlabelled C come from? There is evidence that not all of the starch synthesized during the day reached maximal specific activities based on a comparison of total starch synthesis versus that estimated from the 14C-data (compare Fig. 4A, B). For example, in line 84-9 there was a difference of 50 mmol C m–2 between total starch accumulation during the day (Fig. 4A) and that estimated from 14C-pools (Fig. 4B). Possibly isotopic discrimination is occurring in the starch synthetic pathway. In addition, at the beginning of the 14C labelling there would have been 150 and 34 mmol C m–2 of unlabelled starch in lines 84-9 and 85-1, respectively (Fig. 4A). It is assumed that starch degradation occurs first on the outer layers (newly 14C fixed) of the starch granule. Since there is still some 14C-starch remaining in the leaf at the end of the night, perhaps degradation of the starch occurs concurrently in inner layers (starch from previous days) or new starch granules can be formed at any day. Furthermore, recent studies using 13C labelling have shown that night-time respiration is supported by substrate pools other than newly fixed photosynthates and that starch and sucrose synthesis are fed by recent photosynthates as well as previously fixed low-turnover C compounds (Alonso et al., 2005; Nogués et al., 2004). Other explanations for the low night-time rates estimated by the GM detector is that C is reallocated during the night in different cells layers within the leaf, or transitory changes in leaf thickness or shape occur due to factors such as altered water relations; both possibilities could alter the counting efficiency of the GM counter.
Diel C availability and growth
Extension rates and mass gain of the main stem were higher in 85-1 (Table 2). The wild type showed double the main stem growth during 07.00–15.00 h when export of C was much larger than in 84-9. The stem extension data provides some evidence that at least for the main stem the growth curve can shift towards the daytime when supply of C is greater (53% in 85-1 versus 39% in 84-9 from 07.00–15.00 h, see Table 2). The biggest effect occurred from 07.00–15.00 h when C was the least available to the sinks in 84-9. During this time almost twice as much C was exported in 85-1 and the main stem showed double the growth. From 15.00–23.00 h little difference was seen because, during this period, stem extension was at a minimum in both lines. This indicates that stem growth is not just a direct function of C availability, but providing C when most growth occurs can have a significant effect. These data provide correlative evidence for a relationship between timing of main stem extension growth and C availability. After steady-state 14C export was reached in the day period, the C export rate was relatively equal in the first and second halves of the light period for both lines (Fig. 2B). Therefore, daytime export rates of 108 and 69 mmol C m–2 (calculated from Table 1, assuming that export is the same in the two 8 h light periods) can be estimated between 07.00 h and 15.00 h in lines 85-1 and 84-9, respectively. Correspondingly, during the night period 90 and 109 mmol C m–2 was exported in lines 85-1 and 84-9 (Table 1). Interestingly, main stem growth in 84-9 did not compensate during the night when export was significantly higher compared with the first half of the light period (see above). All of the internode expansion is occurring above the fed leaf, as internodes below have mostly expanded in length and secondary growth is resulting in width increase. As noted above in Fig. 5, during the night there appears to be a preferential allocation of C towards the bottom parts. Thus, in Flaveria plants during the night the C may not be available to the top expanding internodes to allow for a compensation of reduced main stem extension seen in 84-9 during the daytime.
The sucrose supplementation experiment provided further evidence for a relationship between diel timing of C availability and main stem growth. In preliminary experiments, sucrose supplementation through leaves was shown not be an effective method for providing additional sucrose (data not shown). Furbank et al. (1997) showed that Flaveria bidentis can take up sucrose through the roots, and thus it was decided to try this approach. A sterile system was developed that allowed sucrose to be delivered through the roots at specific times of the day. In the present study, sucrose was delivered via the roots between 11.00–12.00 h, which was a time when leaf C export from 84-9 was significantly less compared to 85-1. Maximal effects of sucrose supplementation were seen for line 84-9 during the period just after sucrose application, and stem elongation rates during this period increased to levels found for line 85-1. It is noteworthy that internode elongation rates were affected by sucrose supplementation but not final internode lengths, indicating that internode length in Flaveria is not governed by availability of C. Over an 8–9 d period, sucrose supplementation increased the main stem extension rate for 84-9 to the rate found for 85-1. Currently, the fate of sucrose delivered via the roots in F. linearis is being investigated.
The results of the present study illustrate the importance of considering the relative diel timing of source and sink activities in plants. Circadian rhythms have been shown for net C assimilation in plants (Hennessey and Field, 1991), for photosynthetic gene expression (Harmer et al., 2000), and even spatiotemporal variations in photosynthetic capability have been identified (Rascher et al., 2001). A diel component also exists for C export in plants (Hendrix and Grange, 1991; Geiger and Servaites, 1994; Grodzinski et al., 1999). The present study shows that altered leaf sucrose–starch partitioning due to a reduction in cytFBPase activity can affect the diel patterns of C export. CytFBPase is one logical target for altering diel sucrose synthesis since recent evidence shows that the night-time pathway for sucrose synthesis is cytFBPase-independent (Schleucher et al., 1998). Also, C availability per se is not necessarily the only component of source strength in plants, and processes such as N and S assimilation also need to be considered. In fact, circadian rhythms for genes involved in N and S metabolism have also been identified, and the relative expression patterns of genes involved in C, N, and S assimilation suggest that a co-ordination exists between these processes (Harmer et al., 2000). In conjunction with a diel component to source strength, the growth of plant organs such as vegetative and floral stems, leaves, and fruits and expression patterns for genes involved in cell elongation can also exhibit circadian rhythms (Kristie and Jolliffe, 1985; Fisherman and Genard, 1998; Jouve et al., 1998; Schmundt et al., 1998; Harmer et al., 2000). Harmer et al. (2000) also showed that the timing of circadian gene expression patterns can vary for different genes. Perhaps different plant parts also possess different endogenous diel growth rhythms, and thus altered diel timing of C export and availability could potentially impart differential effects on different plant organs, tissues, and even cells. It is proposed that a consideration of diel patterns are crucial when examining and defining source and sink strength in plants.
Another unresolved question with the F. linearis cytFBPase mutant is how a line with no apparent activity of cytFBPase can still support rates of daytime sucrose synthesis up to 3 µmol C m–2 s–1. One possibility is that PPi-PFK replaces cytFBPase in the hydrolysis of FBP to F6P; however, the reverse reaction is typically favoured in vivo (Stitt et al., 1987). It has been demonstrated here that PPi-PFK activity is present in line 84-9. Bouton et al. (1990) showed that FBP levels are elevated up to 50 times in line 84-9, although F6P levels were not significantly affected. It has been estimated using non-aqueous fractionation techniques that the FBP level normally present in plant cells is around 50 µM (Stitt et al., 1987), and thus a 50 times elevation in this level could bring the FBP concentration up to 2.5 mM. This concentration is sufficient to reverse the direction of the reaction catalysed by PPi-PFK (Yaun et al., 1988). As far as is known, this F. linearis mutant is the only genetic variant in plants identified to date that has null activity for cytFBPase specifically and for any enzyme in the sucrose synthesis pathway. It provides evidence that multiple pathways exist for daytime synthesis of sucrose in plants. It will be worthwhile to determine the nature of this additional pathway(s), and to examine whether this additional pathway(s) is unique to this C3–C4 photosynthetic intermediate or whether the pathway(s) is found across photosynthetic types. In addition, it is realized that unidentified genetic differences in the F. linearis mutant line could be responsible for the observed changes in C partitioning, export, and/or growth. The daytime C-export rate was reduced in the low cytFBPase line as would be predicted, and thus intraspecific genetic variation for diel C export patterns was identified in a natural population. To establish a more definitive relationship between low cytFBPase activity and reduced daytime C export will require the analysis of segregation patterns for these traits using F2 and F3 populations as was done in establishing a relationship between low cytFBPase activity and O2 insensitivity of photosynthesis by Micallef et al. (1996). Work is in progress to complement the cytFBPase mutant using a wild-type version of cytFBPase from line 85-1. In addition, a population segregating for the cytFBPase gene is being examined to determine if a genetic relationship exists between diel C export patterns and developmental patterns of main and lateral stem extension growth. The genetic heterogeneity of this germplasm also allows for the potential identification of interactions between C-export patterns and other traits that have not previously been conceived.
In conclusion, a reduction in cytFBPase activity in 84-9 decreased sucrose synthesis and resulted in lower export and increased starch accumulation during the daytime. Low daytime export in 84-9 was compensated only to a small extent by increased night-time export under the environmental conditions used in the present study. However, low starch accumulation and higher export during the daytime in 85-1 correlated with increased main stem elongation and growth. Sugar supplementation helped to complement the reduced main stem growth rate in line 84-9, and the maximal effects of sugar supplementation were seen during a daily period when C export was significantly reduced in line 84-9. This demonstrates the importance of high export capacity in the light when energy is readily available. Altered sucrose–starch partitioning can affect diel export patterns of sugars from leaves to sinks that can influence plant development and growth including responses such as internode elongation.
|
Acknowledgements |
|---|
Research was supported by grants to BG and BJM from Natural Sciences and Engineering Research Council of Canada, Center for Research in Earth and Space Technology, the Ontario Ministry of Agriculture and Food, CanAdapt Council, Flowers Canada (ON) Inc., and the Ontario Greenhouse Vegetable Growers. The technical assistance of George Lin, Renée Cloutier and Noe Ortiz in the analysis of the 14C-photoassimilates is greatly appreciated, as is technical assistance from Naheed Rana for the analysis of metabolites in the multiple sampling experiment. The authors are also very grateful to Dave Kristie (Department of Biology, Acadia University, Wolfville, Nova Scotia, Canada) for his assistance in the use of rotary motion sensors and associated software.
|
References |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Alonso AP, Vigeolas H, Raymond P, Rolin D, Dieuaide-Noubhani M. 2005. A new substrate cycle in plants. Evidence for a high glucose-phosphate-to-glucose turnover from in vivo steady-state and pulse-labeling experiments with [13C]glucose and [14C]glucose. Plant Physiology 138, 2220–2232.
Bouma TD, De Visser R, Van Leeuwen PH, De Kock MJ, Lambers H. 1995. The respiratory energy requirements involved in nocturnal carbohydrate export from starch-storing mature leaves and their contribution to leaf dark respiration. Journal of Experimental Botany 46, 1185–1194.
Bouton JH, Brown RH, Byrd GT, Sharkey TD. 1990. Inheritance of the reversal of O2 response of photosynthesis in a Flaveria linearis mutant. Plant Physiology 92, 186–190.
Caspar T, Huber SC, Somerville CR. 1985. Alterations in growth, photosynthesis and respiration in a starchless mutant of Arabidopsis thaliana (L.) deficient in chloroplast phosphoglucomutase activity. Plant Physiology 79, 11–17.
Caspar T, Lin T-P, Kakefuda G, Bendow L, Preiss J, Somerville CR. 1991. Mutants of Arabidopsis with altered regulation of starch degradation. Plant Physiology 95, 1181–1188.
Daie J. 1993. Cytosolic fructose-1,6-bisphosphatase: a key enzyme in the sucrose biosynthetic pathway. Photosynthesis Research 38, 5–14.
Dai Z, Ku MSB, Edwards GE. 1996. Oxygen sensitivity of photosynthesis and photorespiration in different photosynthetic types in the genus Flaveria. Planta 198, 563–571.
Edwards GE, Ku MSB. 1987. Biochemistry of C3–C4 intermediates. In: Stumpf PK, Conn EE, eds. The biochemistry of plants, Vol. 10. Hatch MD, Boardman NK, eds. Photosynthesis. London, UK: Academic Press Inc., 275–325.
Fishman S, Genard M. 1998. A biophysical model of fruit growth: simulation of seasonal and diurnal dynamics of mass. Plant, Cell and Environment 21, 739–752.[CrossRef]
Furbank RT, Pritchard J, Jenkins CLD. 1997. Effects of exogenous sucrose feeding on photosynthesis in the C3 plant tobacco and the C4 plant Flaveria bidentis. Australian Journal of Plant Physiology 24, 291–299.
Frommer WB, Sonnewald U. 1995. Molecular analysis of carbon partitioning in solenaceous species. Journal of Experimental Botany 46, 587–607.
Galtier N, Foyer CH, Murchie E, Alred R, Quick P, Voelker TA, Thepenier C, Lasceve G, Betsche T. 1995. Effects of light and atmospheric carbon dioxide enrichment on photosynthesis and carbon partitioning in the leaves of tomato (Lycopersicon esculentum) plants over-expressing sucrose phosphate synthase. Journal of Experimental Botany 46, 1335–1344.
Geiger DR, Fondy BR. 1979. A method for continuous measurement of export from a leaf. Plant Physiology 64, 361–365.
Geiger DR, Servaites JC. 1994. Diurnal regulation of photosynthetic carbon metabolism in C3 plants. Annual Review of Plant Physiology and Plant Molecular Biology 45, 235–256.[CrossRef][Web of Science]
Goodall DW. 1946. The distribution of weight change in the young tomato plants. II. Changes in dry weight of separated organs, and translocation rates. Annals of Botany NS 10, 305–338.
Grodzinski B, Jiao J, Knowles VL, Plaxton WC. 1999. Photosynthesis and carbon partitioning in transgenic tobacco plants deficient in leaf cytosolic pyruvate kinase. Plant Physiology 120, 887–895.
Grodzinski B, Jiao J, Leonardos ED. 1998. Estimating photosynthesis and concurrent export rates in C3 and C4 species at ambient and elevated CO2. Plant Physiology 117, 207–215.
Harmer SL, Hogenesch JB, Straume M, Chang H-S, Han B, Zhu T, Wang X, Kreps JA, Kay SA. 2000. Orchestrated transcription of key pathways in Arabidopsis by the circadian clock. Science 290, 2110–2113.
Hatch MD. 1987. C4 photosynthesis: a unique blend of modified biochemistry, anatomy and ultrastructure. Biochimica et Biophysica Acta 895, 81–106.
Häusler RE, Schlieben NH, Schlulz B, Flügge U-I. 1998. Compensation of decreased triose phosphate/phosphate translocator activity by accelerated starch turnover and glucose transport in transgenic tobacco. Planta 204, 366–376.[CrossRef][Web of Science][Medline]
Hendrix DL, Grange RI. 1991. Carbon partitioning and export from mature cotton leaves. Plant Physiology 95, 228–233.
Hennessey TL, Field CB. 1991. Circadian rhythms in photosynthesis. Plant Physiology 96, 831–836.
Ho LC. 1978. The regulation of carbon transport and the carbon balance of mature tomato leaves. Annals of Botany 42, 155–164.
Jouve L, Greppin H, Agosti RD. 1998. Arabidopsis thaliana floral stem elongation: evidence for an endogenous circadian rhythm. Plant Physiology and Biochemistry 36, 469–472.[CrossRef]
Kehr J, Hustiak F, Walz C, Willmitzer L, Fisahn J. 1998. Transgenic plants changed in carbon allocation pattern display a shift in diurnal growth pattern. The Plant Journal 16, 497–503.[CrossRef][Web of Science][Medline]
Kristie DN, Jolliffe PA. 1985. High-resolution studies of growth oscillations during stem elongation. Canadian Journal of Botany 64, 2399–2405.
Ku SBM, Wu J, Dai Z, Scott RA, Chun C, Edwards GE. 1991. Photosynthetic and photorespiratory characteristics of Flaveria species. Plant Physiology 96, 518–528.
Leonardos ED, Grodzinski B. 2000. Photosynthesis, immediate export and carbon partitioning in source leaves of C3, C3-C4 intermediate and C4 Panicum and Flaveria species at ambient and elevated CO2 levels. Plant, Cell and Environment 23, 839–851.
Leonardos ED, Grodzinski B. 2001. Quantifying immediate C export from source leaves. In: Pesarakli M, ed. Handbook of plant and crop physiology, 2nd edn. New York, NY: Marcel Dekker Inc., 407–420.
Leonardos ED, Grodzinski B. 2003. Export patterns of 14C-assimilates from source leaves of C3, C3–C4 intermediate, and C4 Panicum and Flaveria species during light and dark periods. Canadian Journal of Botany 81, 464–476.
Leonardos ED, Savitch LV, Huner NPA, Oquist G, Grodzinski B. 2003. Daily photosynthetic and C-export patterns in winter wheat leaves during cold stress and acclimation. Physiologia Plantarum 117, 521–531.[CrossRef][Medline]
Leonardos ED, Tsujita MJ, Grodzinski B. 1996. The effect of source or sink temperature on photosynthesis and 14C-partitioning in and export from a source leaf in Alstroemeria. Physiologia Plantarum 97, 563–575.[CrossRef]
Lin T-P, Caspar T, Somerville C, Preiss J. 1988. Isolation and characterisation of a starchless mutant of Arabidopsis thaliana (L.) Heynh. lacking ADP-glucose pyrophosphorylase activity. Plant Physiology 86, 1131–1135.
Micallef BJ, Haskins KA, Vanderveer PJ, Roh K-S, Shewmaker CK, Sharkey TD. 1995. Altered photosynthesis, flowering, and fruiting in transgenic tomato plants that have an increased capacity for sucrose synthesis. Planta 196, 327–334.
Micallef BJ, Sharkey TD. 1996. Genetic and physiological characterization of Flaveria linearis plants having a reduced activity of cytosolic fructose-1,6-bisphosphatase Plant, Cell and Environment 19, 1–9.
Micallef BJ, Vanderveer PJ, Sharkey TD. 1996. Responses to elevated CO2 of Flaveria linearis having a reduced activity of cytosolicfructose-1,6-bisphosphatase. Plant, Cell and Environment 19, 10–16.
Monson RK, Moore Bd, Ku MSB, Edwards GE. 1986. Co-function of C3- and C4-photosynthetic pathways in C3, C4, and C3–C4 intermediate Flaveria species. Planta 168, 493–502.[CrossRef]
Moore B, Zhou L, Rolland F, Hall Q, Cheng W-H, Liu Y-X, Hwang I, Jones T, Sheen J. 2003. Role of the Arabidopsis glucose sensor HXK1 in nutrient, light, and hormonal signaling. Science 300, 332–336.
Nielsen TH, Stitt M. 2001. Tobacco transformants with strongly decreased expression of pyrophosphate: fructose-6-phosphate expression in the base of their young growing leaves contain much higher levels of fructose-2,6-bisphosphate but no major changes in fluxes. Planta 214, 106–116.[Web of Science][Medline]
Noguchi K, Go C-S, Miyazawa S-I, Terashima I, Ueda S, Yoshinari T. 2001. Costs of protein and carbohydrate export in leaves of sun and shade species. Australian Journal of Plant Physiology 28, 37–47.[Web of Science]
Nogués S, Tcherkez G, Cornic G, Ghashghaie J. 2004. Respiratory carbon metabolism following illumination in intact French bean leaves using 13C/12C isotope labelling. Plant Physiology 136, 3245–3254.
Rascher U, Hutt M-T, Siebke K, Osmond B, Beck F, Luttge U. 2001. Spatiotemporal variation of metabolism in a plant circadian rhythm: The biological clock as an assembly of coupled individual oscillators. Proceedings of the National Academy of Sciences, USA 98, 11801–11805.
Riesmeier JW, Flugge UI, Schlulz B, Heineke D, Heldt HW, Willmitzer L, Frommer WB. 1993. Antisense repression of the chloroplast triose phosphate translocator affects carbon partitioning in transgenic potato plants. Proceedings of the National Academy of Sciences, USA 90, 6160–6164.
Schleucher J, Vanderveer PJ, Sharkey TD. 1998. Export of carbon from chloroplasts at night. Plant Physiology 118, 1439–1445.
Schmundt D, Stitt M, Jahne B, Schurr U. 1998. Quantitative analysis of the local rates of growth of dicot leaves at high temporal and spatial resolution, using image sequence analysis. The Plant Journal 16, 505–514.[CrossRef][Web of Science]
Sharkey TD, Savitch LV, Vanderveer PJ, Micallef BJ. 1992. Carbon partitioning in a Flaveria linearis with reduced cytosolic fructose bisphosphatase. Plant Physiology 100, 210–215.
Sheen J, Zhou L, Jang JC. 1999. Sugars as signaling molecules. Current Opinion on Plant Biology 2, 410–418.
Stitt M, Huber S, Kerr P. 1987. Control of photosynthetic sucrose formation In: Stumpf PK, Conn EE, eds. The biochemistry of plants, Vol. 10. Hatch MD, Boardman NK, eds. Photosynthesis. London, UK: Academic Press Inc., 328–409.
Stitt M, Sonnewald U. 1995. Regulation of metabolism in transgenic plants. Annual Review of Plant Physiology and Plant Molecular Biology 46, 341–368.[CrossRef][Web of Science]
Strand A, Zrenner R, Trevanion S, Stitt M, Gustafsson P, Gardestrom P. 2000. Decreased expression of two key enzymes in the sucrose biosynthesis pathway, cytosolic fructose-1,6-bisphosphatase and sucrose phosphate synthase, has remarkably different consequences for photosynthetic carbon metabolism in transgenic Arabidopsis thaliana. The Plant Journal 23, 759–770.[CrossRef][Web of Science][Medline]
Strand A, Foyer CH, Gustafsson P, Gardestrom P, Hurry V. 2003. Altering flux through the sucrose biosynthesis pathway in transgenic Arabidopsis thaliana modifies photosynthetic acclimation at low temperatures and the development of freezing tolerance. Plant, Cell and Environment 26, 523–535.[CrossRef]
Terry N, Mortimer DC. 1972. Estimation of the rates of mass carbon transfer by leaves of sugar beet. Canadian Journal of Botany 50, 1049–1054.
Yaun X-H, Kwiatkowska D, Kemp RG. 1988. Inorganic pyrophosphate: fructose-6-phosphate 1-phosphotransferase of the potato tuber is related to the major ATP-dependent phosphofructokinase of E. coli. Biochemical and Biophysical Research Communications 154, 113–117.[CrossRef][Web of Science][Medline]
Zeeman SC, ap Rees T. 1999. Changes in carbohydrate metabolism and assimilate export in starch-excess mutants of Arabidipsis. Plant, Cell and Environment 22, 1445–1453.[CrossRef]
Zeeman SC, Northrop F, Smith AM, ap Rees T. 1998. A starch-accumulating mutant of Arabidipsis thaliana deficient in a starch-hydrolysing enzyme. The Plant Journal 15, 357–365.[CrossRef][Web of Science][Medline]
Zrenner R, Krause KP, Apel P, Sonnewald U. 1996. Reduction of the cytosolic fructose-1,6-bisphosphatase in transgenic potato plants limits photosynthetic sucrose biosynthesis with no impact on plant growth and tuber yield. The Plant Journal 9, 671–681.[CrossRef][Web of Science][Medline]
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  A. J. Serrato, J. de Dios Barajas-Lopez, A. Chueca, and M. Sahrawy Changing sugar partitioning in FBPase-manipulated plants J. Exp. Bot., March 26, 2009; (2009) erp066v1. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||