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Plant Physiol. (1998) 116: 1315-1322
Cyst(e)ine Is the Transport Metabolite of Assimilated Sulfur from
Bundle-Sheath to Mesophyll Cells in Maize Leaves1
Marta Burgener,
Marianne Suter,
Stephanie Jones, and
Christian Brunold*
Institute of Plant Physiology, Altenbergrain 21, CH-3013 Berne,
Switzerland
 |
ABSTRACT |
The intercellular distribution of the
enzymes and metabolites of assimilatory sulfate reduction and
glutathione synthesis was analyzed in maize (Zea mays L. cv LG 9) leaves. Mesophyll cells and strands of bundle-sheath cells
from second leaves of 11-d-old maize seedlings were obtained by two
different mechanical-isolation methods. Cross-contamination of cell
preparations was determined using ribulose bisphosphate carboxylase (EC
4.1.1.39) and nitrate reductase (EC 1.6.6.1) as marker enzymes for
bundle-sheath and mesophyll cells, respectively. ATP sulfurylase (EC
2.7.7.4) and adenosine 5 -phosphosulfate sulfotransferase activities
were detected almost exclusively in the bundle-sheath cells, whereas
GSH synthetase (EC 6.3.2.3) and cyst(e)ine,  -glutamylcysteine, and
glutathione were located predominantly in the mesophyll cells. Feeding
experiments using [35S]sulfate with intact leaves
indicated that cyst(e)ine was the transport metabolite of reduced
sulfur from bundle-sheath to mesophyll cells. This result was
corroborated by tracer experiments, which showed that isolated
bundle-sheath strands fed with [35S]sulfate secreted
radioactive cyst(e)ine as the sole thiol into the resuspending medium.
The results presented in this paper show that assimilatory sulfate
reduction is restricted to the bundle-sheath cells, whereas the
formation of glutathione takes place predominantly in the mesophyll
cells, with cyst(e)ine functioning as a transport metabolite between
the two cell types.
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INTRODUCTION |
Maize (Zea mays L.), as a typical
C4 plant, is characterized by a compartmentation
of carbon assimilation into specific cell types, with
CO2 initially being fixed into malate in the MC
and then transported into the BSC, where the formation of glycerate 3-phosphate is localized (Black, 1973 ). Conversely, the reduction of
nitrate occurs exclusively in the MC (Moore and Black, 1979). This
division of labor is a primary factor contributing to high rates of
carbon assimilation (Black, 1973) and nitrogen use efficiency (Brown,
1978) in C4 plants.
The intercellular compartmentation of sulfate assimilation is less well
documented. Sulfite reductase (EC 1.8.7.1) and
O-acetyl-l-Ser sulfhydrylase (EC 4.2.99.8)
activities were found in both cell types at comparable levels (Passera
and Ghisi, 1982; Burnell, 1984; Schmutz and Brunold, 1984, 1985).
ATPSase (EC 2.7.7.4), the first enzyme in the pathway of assimilatory
sulfate reduction, is, however, reported to be predominantly or even
exclusively located in the BSC (Gerwick and Black, 1979; Gerwick et
al., 1980; Passera and Ghisi, 1982; Burnell, 1984; Schmutz and Brunold,
1984; Ghisi et al., 1986). In addition, the second step in sulfate
reduction, catalyzed by APSSTase (Brunold and Rennenberg, 1997), an
enzyme that may be identical to 5-adenylylsulfate reductase recently described by Setya et al. (1996), is essentially restricted to the BSC
(Schmutz and Brunold, 1984).
There are no published results concerning the intercellular
compartmentation of GSH synthesis. This tripeptide plays an important role in the plant's defense system (Rennenberg and Brunold, 1994). It
is involved in various stress situations, such as heavy metal stress
(Nussbaum et al., 1988; Rüegsegger and Brunold, 1992; Galli et
al., 1996), xenobiotic stress (Farago et al., 1994), and chilling
responses (Kocsy et al., 1996). GSH is also an essential factor in the
regulation of sulfur nutrition in plants (Brunold and Rennenberg,
1997). Its long-distance transport mediates distribution of reduced
sulfur according to the requirements of individual plant organs, and
controls sulfate influx into the plant (Rennenberg and Lamoureux, 1990;
Herschbach and Rennenberg, 1994; Lappartient and Touraine, 1996). There
is also evidence for membrane transport of GSH in an active,
carrier-mediated process (Rennenberg and Lamoureux, 1990). These facts,
together with the localization of enzymes involved in assimilatory
sulfate reduction, raise the question of the nature of thiol compounds
transported from BSC to MC in C4
plants.
In this paper we present evidence that cyst(e)ine is the transport
metabolite and that GSH synthesis takes place predominantly in MC.
| |
MATERIALS AND METHODS |
Fifty maize (Zea mays L. cv LG 9, Limagrain, Ennezat,
France) kernels were soaked for 24 h in aerated tap water at room
temperature and then transferred to a pot containing 5000 cm3 of moist Perlite (Samen Mauser, Berne,
Switzerland). After 3 d in the dark, the seedlings were cultivated
in a 16-/8-h photoperiod at 25/20°C with a PPFD of 350 µmol
m 2 s1 and 70% RH. One
liter of nutrient solution (Henschel, 1970), modified according to the
method of Nussbaum et al. (1988), was added every 3rd d. Second leaves
were harvested 10 d after imbibition in the middle of the light
period.
Cell Isolation
MC and BSC extracts were obtained by two different
mechanical-isolation methods.
Method A Modified after Schmutz and Brunold (1984)
One gram of leaves was cut vertically across the veins in
approximately 1-mm segments with a razor blade and transferred to 10 mL
of medium containing 2% cellulase and 2% pectinase for protoplast isolation, according to the method of Mills and Joy (1980). The plant
material was infiltrated by applying a vacuum and then incubated for 20 min at 30°C. The predigested leaf strips were thoroughly rinsed and
then homogenized in 8 mL of 0.1 m Tris-HCl (pH 8.0) containing 20 mm MgCl, 100 mm KCl, and 10 mm dithioerythritol. The extraction was carried out using a
Sorvall Omni Mixer for 10 s at 100 V (6,700 rpm) and twice for
15 s at 170 V (11,500 rpm). The homogenate was filtered through a
60-µm nylon net. The filtrate contained the broken MC. The plant
material on the nylon net was resuspended in 8 mL of extraction buffer
and homogenized again twice for 20 s at 240 V (16,250 rpm), and
was then filtered and rinsed. The bundle-sheath strands on the nylon
net were collected and broken in 4 mL of extraction buffer using a
cooled glass homogenizer.
Method B Using a Roller Device according to the Method of
Leegood (1985)
The harvested leaves were cut into 3- to 4-cm sections. Leaf
segments, placed on 100 µL of extraction buffer, were rolled once,
thus squeezing out MC. The sap from two leaves was collected with a
pipette and resuspended in 300 µL of extraction buffer. The rolled
leaf laminas were extracted with a cooled glass homogenizer in 4 mL of
extraction buffer. The activities of these extracts as well as their
thiol content were corrected for pure BSC given that NR is restricted
to MC (Moore and Black, 1979; Vaughn and Campbell, 1988).
All extracts were filtered through two layers of 100% viscose fleece
(Milette, Migros, Switzerland). Cross-contamination of the cell
preparations was determined using Rubisco and NR as marker enzymes for
BSC and MC, respectively.
Enzyme Assays
Rubisco activity was determined according to the method of
Buchanan and Schürmann (1973) by measuring the nonvolatile
radioactivity produced from RuBP and
H14CO3
and by following the modifications given by Wyss and Brunold (1979).
Incubation was for 10 min at 30°C. NR measurement was carried out
according to the method of Neyra and Hagemann (1975) with modifications
described by Kast et al. (1995). ATPSase activity was determined by
measuring production of ATP from APS and PPi with a
luciferin-luciferase system (Schmutz and Brunold, 1982) using a
Lumac/3M Biocounter (model M 2010, Lumac, Basel, Switzerland). APSSTase
activity was measured as the production of
[35S]sulfite, assayed as acid volatile
radioactivity formed from [35S]APS (Brunold and
Suter, 1990) in the presence of dithioerythritol. GSH synthetase
activity was measured as previously reported (Klapheck et al., 1987;
Rüegsegger and Brunold, 1992) by the quantification of the
reaction product after reverse-phase HPLC separation of its
monobromobimane derivative.
The activities of Rubisco, NR, APSSTase, and GSH synthetase were
determined immediately after extraction. ATPSase activity, which was
stable in the crude extract up to at least 6 h (data not shown),
was routinely measured 1 h after the homogenization of the plant
material.
Determination of RuBP
RuBP was determined by the incorporation of
14CO2 into an acid-stable
product as described by Doulis et al. (1997).
Determination of Cyst(e)ine, EC, and GSH
Thiols were separated and quantified by reverse-phase HPLC after
reduction with NaBH4 and fluorescent labeling
with monobromobimane (Newton et al., 1981; Schupp and Rennenberg, 1988)
as previously described (Rüegsegger and Brunold, 1992).
Recoveries of 88, 90, and 98% were determined for cyst(e)ine, EC,
and GSH, respectively, and were used for calculating the actual amounts
of the thiols in the plant material. The measured values were related
to average protein content of extracts.
Tracer Experiment
For [35S]sulfate labeling, second leaves
were excised and placed into 0.5 mL of nutrient solution containing 75 instead of 750 µm sulfate and 5.55 × 106 Bq of [35S]sulfate.
The incubation took place under cultivation conditions. At different
times, leaves were immediately extracted with isolation method B. The
sections that had been in contact with the nutrient solution were
excised and discarded. The experiments were repeated with the method
described for determination of thiols, including the acidic extraction.
Measurement of 35S Label in Thiols and Sulfide
The determination of the radioactive sulfide content required an
alkaline-extraction method. The extraction buffer consisted of 200 mm 2-(cyclohexyl-amino)-ethanesulfonic acid-NaOH (pH 8.4) containing 1 mm Na2EDTA. Three
hundred microliters of extract was reduced on ice with 30 µL of
freshly prepared NaBH4 for 30 min. For
derivatization, 220 µL of this mixture was added to 20 µL of 15 mm monobromobimane and kept in the dark at room temperature for 15 min. The reaction was stopped with 165 µL of 5% (v/v) acetic acid.
A 50-µL aliquot of each sample was separated by HPLC, as described
for the inactive thiols, using a modified gradient of methanol (0-14%
methanol in 15 min and then 14-51% methanol in 30 min). Fractions of
0.75 mL were collected in scintillation vials. Two milliliters of
Ultima Gold XR scintillation cocktail (Packard, Zürich, Switzerland)
was added per fraction, and the radioactivity was counted in a
Betamatic V liquid-scintillation counter (Kontron, Zürich,
Switzerland).
35S Labeling of Isolated Bundle-Sheath
Strands
Two grams of 1-mm leaf segments was blended in 16 mL of medium
according to the method of Valle and Heldt (1991) in a polytron with
1-s (20,000 rpm) and 20-s (15,000 rpm) bursts. The bundle-sheath strands were collected on a 280-µm nylon net, washed thoroughly, and
resuspended at a level of approximately 50 µg chlorophyll mL1 resuspending medium (0.3 m
sorbitol, 20 mm Tricine-KOH (pH 7.5), 4 mm
MgCl2, 10 mm KCl, 2 mm
KH2PO4, 1 mm
ADP, 10 mm glycerate 3-phosphate, 5 mm
glutamate, 5 mm malate, 2 mm
O-acetyl-l-Ser, and 0.2 mm
K2SO4). The bundle-sheath
strands were incubated after the addition of 1.85 × 107 Bq [35S]sulfate
mL1 at 30°C with an irradiance of
approximately 1500 µmol m2
s1. At different times, supernatants of the
suspended bundle-sheath strands were obtained by a 5-min centrifugation
at 10,000 rpm at 4°C. The supernatant was treated with 0.3 mm DTT for reducing possibly existing thiols, derivatized,
and separated by HPLC as described for cyst(e)ine, EC, and GSH. The
injection volume was 100 µL. Fractions of 0.375 mL were collected and
determined as in the tracer experiment with second leaves.
The incubated BSC were washed extensively and extracted in 0.1 n HCl. The determination of
35S-labeled substances was performed as
described for the supernatant.
Protein Determination
The protein content of the extracts was measured according to the
method of Bradford (1976) with BSA as the standard.
Chlorophyll Determination
Acetone was added to the bundle-sheath strands and the mixture was
shaken several times for 30 min in the dark. Chlorophyll was measured
in the clear supernatant after centrifugation using the procedure of
MacKinney (1941).
Statistical Analysis
The Mann-Whitney rank sum test (SigmaStat for Windows, version
1.0, 1992-1994, Jandel, San Rafael, CA) was used to determine significant differences in the enzyme activities and concentrations of
metabolites between extracts of BSC and MC.
Chemicals
Monobromobimane was obtained from Calbiochem and EC was
obtained from Nacalai Tesque (Kyoto, Japan).
[35S]APS was prepared according to the method
of Li and Schiff (1991), using ATPSase from Sigma and
[35S]sulfate from the Radiochemical Centre
(Amersham). All other chemicals were purchased from Fluka.
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RESULTS |
Rubisco was assayed as a marker for BSC and NR was assayed as a
marker for MC to determine the level of cross-contamination in the cell
preparations. The activity of Rubisco indicated that there was less
than 10% of BSC in the mesophyll extracts obtained by method A (Fig.
1A) and less than 5% in those obtained
with the roller device (Fig. 1B). Based on the distribution of NR
activity, procedure A yielded very pure BSC extracts with a
contamination of about 5% (Fig. 1A), whereas the rolled leaf laminas
in method B still contained an appreciable amount of MC (Fig. 1B).
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| Figure 1.
A, Intercellular localization of activities of
Rubisco (first panel), NR (second panel), APSSTase (third panel), and
ATPSase (fourth panel) in second leaves of 11-d-old maize plants. MC
(open bars) and BSC (black bars) extracts were obtained by isolation procedure A. Mean values ± sd of four different
experiments are presented. The values of the extracts of the two cell
types differ significantly from each other (P  0.05). B,
Intercellular localization of activities of Rubisco (top), NR (middle),
and GSH synthetase (bottom) in MC (open bars) and BSC (black bars)
extracts obtained by isolation method B. Means ± sd
of five independent experiments are presented. The values of the
extracts of the two cell types differ significantly from each other
(P 0.05).
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The distribution of ATPSase and APSSTase (the two first enzymes of
assimilatory sulfate reduction) between BSC and MC is presented in
Figure 1A. Both are predominantly located in BSC. The APSSTase activity
in the MC extracts was at the same relative level as Rubisco activity,
indicating that this enzyme is exclusively active in the BSC. The
relative level of ATPSase activity detected in the MC extract is higher
than expected from contaminating BSC, indicating that this enzyme of
sulfate assimilation is active in both cell types, with more than 90%
of total activity in BSC.
Figures 1B and 2
show the compartmentation of GSH synthetase and thiols. After
correction for contamination, only 22% of the total GSH synthetase
activity was located in BSC (Fig. 1B). Also, the amounts of the thiols
cyst(e)ine, EC, and GSH in the BSC extracts were significantly lower
compared with those in the MC preparations (Fig. 2). The content of
RuBP in the MC extract was used to calculate the degree of
contamination by low-Mr compounds originating in
the BSC (Doulis et al., 1997). The RuBP distribution between extracts
of BSC and MC was similar to that of Rubisco (data not shown),
indicating that the concentrations of the thiols were not a consequence
of leaky plasma membranes.
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| Figure 2.
Contents of cyst(e)ine (top), EC (middle), and
GSH (bottom) in BSC (black bars) and MC (open bars). Cell types were
obtained by isolation procedure B. The values of BSC were corrected for contamination by MC. Means ± sd of four independent
experiments are presented. The values of the two cell types differ
significantly from each other (P 0.05).
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Mixing of extracts from BSC and MC gave additive activities of all
measured enzymes, indicating that no inactivator was present in either
cell type and that the activities were comparable to those of the whole
leaves, showing that no activity was lost during cell separation (data
not shown).
The distribution of the activities of the two first enzymes of
assimilatory sulfate reduction indicated an almost exclusive localization of this pathway in BSC, whereas GSH synthesis
predominantly took place in MC. This opened the question of
intermediates being synthesized in BSC and transported into MC. We
addressed this question by feeding excised leaves with
[35S]sulfate and analyzing the radioactivity of
the thiols in the MC in a time-course experiment. The percentage of
35S-radiolabeled thiols in MC extracts from such
an experiment is presented in Figure
3A. Seventy-five
percent of the radioactivity of the thiols in MC was detected in
cyst(e)ine after a 5-min incubation, whereas after 90 min the label was
found at a comparable percentage in GSH. The recovery of cyst(e)ine,
EC, and GSH was 36 to 37% in comparison with the acidic procedure
as a standard method for thiol extraction (data not shown). Only
negligible amounts of [35S]sulfide were
detected at all times, although the recovery rate of sulfide with the
alkaline-extraction method was as high as those of the thiols at any
time. The repetition of the experiment, including the acidic
extraction, resulted in the same time course of radioactivity in the
thiols (Fig. 3B). The amount of 35S-radiolabeled
thiols in the BSC during the incubation time was very low compared with
those in MC (Fig. 3C). After a 90-min incubation, 75% of the total
radioactivity in the thiols was measured in GSH of MC. Calculated on
the basis of the specific activity of
[35S]sulfate fed to the leaves, this
percentage corresponded to 0.13 nmol GSH mg1
protein.
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| Figure 3.
A, Radioactivity in cyst(e)ine ( ), EC
( ), and GSH ( ) in MC as a percentage of total radioactivity in
the three compounds after an alkaline-extraction method. Radioactivity
was supplied as [35S]sulfate by incubating 11-d-old
second leaves for 5 to 90 min. Values of two independent experiments at
each time are presented. B, Radioactivity in cyst(e)ine (), EC
(), and GSH () in MC as a percentage of total radioactivity in
the three compounds after an acid-extraction method. Means ± sd of four independent experiments at each time are
presented. C, Radioactivity in cyst(e)ine (top), EC (middle), and
GSH (bottom) in BSC (left) and MC (right) as a percentage of the sum of
the three compounds in both cell types after acidic extraction. The
values of BSC were corrected for contamination by MC. Mean values ± sd of four independent experiments at each time are
presented.
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Taken together with the distribution of the enzyme activities involved,
these results indicate that cyst(e)ine is the thiol transported from
BSC to MC, where it is used for GSH synthesis. To corroborate this
result, we incubated isolated bundle-sheath strands in a medium
containing [35S]sulfate. These experiments
resulted in only one radioactive product detectable in the resuspending
medium. Figure 4 shows the HPLC
chromatographs of 35S-labeled substances in the
resuspending medium for different incubation times. The peak with a
retention time of 2.5 min represents the supplied
[35S]sulfate, and the signal at 13.5 min
resulted from the washing of the column. The only other substance
detected with appreciable radioactivity was identified as cyst(e)ine by
injecting monobromobimane derivatives of Cys, EC, and GSH and
measuring them with a fluorescence detector. Calculated on the basis of
the specific activity of [35S]sulfate fed to
the isolated BSC, the amounts of synthesized cyst(e)ine in the 15- and
30-min incubations were 1.2 and 2.4 nmol mg1
chlorophyll, respectively. The small peak at the beginning of incubation corresponding to 0.5 nmol cyst(e)ine
mg1 chlorophyll can be most likely explained by
synthesis of the product during separation of BSC from resuspending
medium. In a parallel experiment, in which the BSC were centrifuged
first and the resulting supernatant was incubated with
[35S]sulfate, no cyst(e)ine was detectable
(data not shown).
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| Figure 4.
Separation of 35S compounds in the
incubation medium of isolated bundle-sheath strands by HPLC. Incubation
with [35S]sulfate was for 0, 15, and 30 min. The arrow
indicates the retention time of the monobromobimane derivative of
cyst(e)ine.
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To exclude the possibility that the above-mentioned results were
due to leaky plasma membranes we measured RuBP as a marker (Doulis et
al., 1997). The similar amounts of RuBP that were detected in BSC
before and after the incubation indicated that the chloroplasts and
plasma membranes of these cells were not leaky for all
low-Mr molecules caused by the exposure (data
not shown).
| |
DISCUSSION |
Because of their lower stability, MC disrupt preferentially in a
mechanical isolation procedure. This characteristic makes it possible
to obtain very pure MC extracts within seconds using a roller device
(Leegood, 1985). The rapid extraction method is, therefore, excellent
for short-time tracer experiments and measurements of enzyme activities
in MC. If pure extracts of both MC and BSC are of importance, then the
leaves should be predigested in a medium for protoplast isolation
(Mills and Joy, 1980) before the cell types are separated mechanically
(Schmutz and Brunold, 1984).
Bundle-sheath strands consisting of a segment of vascular bundle
surrounded by BSC of high functional integrity and metabolic competence
can be obtained easily by blending leaf segments in an appropriate
medium with a polytron (Valle and Heldt, 1991). The intact
plasmodesmata that originally connected the bundle-sheath cytosol with
MC are permeable to molecules up to a molecular mass of about 900 D
(Valle et al., 1989). This means that the highly intact BSC are
accessible to substrates added to the resuspending medium, thus
enabling metabolic studies.
The findings presented in this paper suggest a cooperation between BSC
and MC in sulfate reduction and GSH synthesis, as shown in Figure
5. The scheme compares the intercellular
localization of sulfur assimilation with the compartmentation of
reactions involved in carbon and nitrogen assimilation. The
distribution of ATPSase and APSSTase activities in maize was
consistent with previous results, which indicated a preponderant or
even exclusive localization of assimilatory sulfate reduction to BSC
(Burnell, 1984; Schmutz and Brunold, 1984, 1985; Ghisi et al., 1986).
The high percentage of GSH synthetase activity in MC indicates that this cell type is the main site for synthesis of the tripeptide in
maize leaves and correlates with the high levels of GSH determined in
MC, as compared with BSC.
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| Figure 5.
Diagram of the hypothetical intercellular
distribution of CO2, nitrate, and sulfate assimilation and
GSH synthesis in maize leaves. Cys and Gly exported from BSC and Glu
formed in MC are used for GSH synthesis. VB, Vascular
bundle.
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Information about intermediates of sulfate assimilation transported
from BSC to MC was provided by feeding intact leaves with [35S]sulfate and then rapidly extracting the
MC. The immediate appearance of 35S-labeled
cyst(e)ine in the MC indicated that this amino acid is the molecule
transporting reduced sulfur. This hypothesis was corroborated by the
finding that isolated bundle-sheath strands released cyst(e)ine into
the resuspending medium. Since no cyst(e)ine was detectable in a
parallel experiment in which the BSC were centrifuged first, and only
the resulting supernatant of the suspension was incubated with
[35S]sulfate, we can be sure that the
[35S]cyst(e)ine was synthesized in the
intact BSC and then transported into the resuspension medium. In a
physiological context this cyst(e)ine would be exported into MC.
The compartmentation of CO2 and nitrate
assimilation in C4 plants leads to important
advantages with regard to carbon (Black, 1973) and nitrogen use
efficiency (Brown, 1978). The physiological reason for the
cell-type-specific localization of sulfate assimilation and GSH
synthesis between BSC and MC is not clear. Sulfate assimilation is
located in BSC. These enclose the vascular bundles, which are the
source of sulfate originating from the soil. It is plausible that
reduced levels of PSII (Sheen and Bogorad, 1988; Pfündel et al.,
1996) and correspondingly lower O2 concentrations
in BSC compared with MC could prevent autooxidation of the reaction
intermediates of assimilatory sulfate reduction, i.e. sulfite and/or
sulfide. A second reason might be that Gly decarboxylase and Ser
hydroxymethyl transferase (enzymes synthesizing Ser, the precursor of
Cys) are localized exclusively in BSC of C4
plants (Gardeström et al., 1978; Ohnishi and Kanai, 1983; Becker
et al., 1993). Since Gly is also produced in this cell type (Martin et
al., 1983; Farineau et al., 1984; Yamaya and Oaks, 1988), GSH synthesis
in the MC could proceed using Cys and Gly formed in BSC and Glu from MC (Fig. 5). A predominant localization of GSH synthesis and
correspondingly high levels of GSH in MC would be advantageous for the
plant to react against reactive oxygen species (Foyer et al., 1994;
Doulis et al., 1997), which are probably produced preferentially in
this cell type in various stress situations (Rennenberg and Brunold, 1994) and during pathogen attack (Low and Merida, 1996).
| |
FOOTNOTES |
1
This work was supported by the Swiss National
Science Foundation. The term "Cys" is used when it is clear that
cystine is not involved; "cyst(e)ine" is used for an undefined
mixture of Cys and cystine. The concentrations are expressed in all
cases relative to Cys.
*
Corresponding author; e-mail chbrunold{at}pfp.unibe.ch; fax
41-31-332-20-59.
Received August 18, 1997;
accepted December 19, 1997.
| |
ABBREVIATIONS |
Abbreviations:
APS, adenosine 5-phosphosulfate.
APSSTase, adenosine 5-phosphosulfate sulfotransferase.
ATPSase, adenosine
triphosphate sulfurylase.
BSC, bundle-sheath cell(s).
EC, -glutamylcysteine.
MC, mesophyll cell(s).
NR, nitrate reductase.
RuBP, ribulose-1,5-bisphosphate.
| |
ACKNOWLEDGMENTS |
We would like to thank Dr. Stanislav Kopriva and Thomas Imhasly
for helpful discussions and Dr. Andrew Fleming for improving the style
of the manuscript.
| |
LITERATURE CITED |
Becker TW,
Perrot-Rechenmann C,
Suzuki A,
Hirel B
(1993)
Subcellular and immunocytochemical localization of the enzymes involved in ammonia assimilation in mesophyll and bundle-sheath cells of maize leaves.
Planta
191:
129-136
Black CC
(1973)
Photosynthetic carbon fixation in relation to net CO2 uptake.
Annu Rev Plant Physiol
24:
253-286
Bradford MM
(1976)
A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye-binding.
Anal Biochem
72:
248-254
[CrossRef][ISI][Medline]
Brown RH
(1978)
A difference in N use efficiency in C3 and C4 plants and its implications in adaptation and evolution.
Crop Sci
18:
93-98
Brunold C,
Rennenberg H
(1997)
Regulation of sulfur metabolism in plants: first molecular approaches.
Prog Bot
58:
164-186
Brunold C,
Suter M
(1990)
Adenosine 5-phosphosulfate sulfotransferase.
Methods Plant Biochem
3:
339-343
Buchanan BB,
Schürmann P
(1973)
Regulation of ribulose 1,5-diphosphate carboxylase in the photosynthetic assimilation of carbon dioxide.
J Biol Chem
248:
4956-4964
[Abstract/Free Full Text]
Burnell JN
(1984)
Sulfate assimilation in C4 plants.
Plant Physiol
75:
873-875
[Abstract/Free Full Text]
Doulis AG,
Debian N,
Kingston-Smith AH,
Foyer CH
(1997)
Differential localization of antioxidants in maize leaves.
Plant Physiol
114:
1031-1037
[Abstract]
Farago S,
Brunold C,
Kreuz K
(1994)
Herbicide safeners and glutathione metabolism.
Physiol Plant
91:
537-542
[CrossRef]
Farineau J,
Lelandais M,
Morot-Gaudry JF
(1984)
Physiol Plant
60:
208-214
Foyer CH,
Lelandais M,
Kunert KJ
(1994)
Photooxidative stress in plants.
Physiol Plant
92:
696-717
[CrossRef]
Galli U,
Schüepp H,
Brunold C
(1996)
Thiols in cadmium- and copper-treated maize (Zea mays L.).
Planta
198:
139-143
Gardeström P,
Edwards GE,
Henricson D,
Ericson I
(1978)
The localization of serine hydroxymethyl-transferase in leaves of C3 and C4 species.
Plant Cell Physiol
19:
1399-1405
[Abstract/Free Full Text]
Gerwick BC,
Black CC
(1979)
Sulfur assimilation in C4 plants.
Plant Physiol
64:
590-593
[Abstract/Free Full Text]
Gerwick BC,
Ku SB,
Black CC
(1980)
Initiation of sulfate activation: a variation in C4 photosynthesis plants.
Science
209:
513-515
[Abstract/Free Full Text]
Ghisi R,
Anaclerio F,
Passera C
(1986)
Effects of nitrogen deprivation on the ATP-sulphurylase and O-acetylserine sulphydrylase activities of mesophyll protoplasts and bundle sheath strands of maize leaves.
Biol Plant
28:
114-119
Henschel G (1970) Untersuchungen über die Aufnahme von
15N-markiertem Harnstoff bei Phaseolus
vulgaris L. PhD thesis. University of Hohenheim, Stuttgart,
Germany
Herschbach C,
Rennenberg H
(1994)
Influence of glutathione (GSH) on net uptake of sulphate and sulphate transport in tobacco plants.
J Exp Bot
45:
1069-1076
[Abstract/Free Full Text]
Kast D,
Stalder M,
Rüegsegger A,
Galli U,
Brunold C
(1995)
Effects of NO2 and nitrate on sulfate assimilation in maize.
J Plant Physiol
147:
9-14
Klapheck S,
Latus C,
Bergmann L
(1987)
Localization of glutathione synthetase and distribution of glutathione in leaf cells of Pisum sativum L.
J Plant Physiol
131:
123-131
Kocsy G,
Brunner M,
Rüegsegger A,
Stamp P,
Brunold C
(1996)
Glutathione synthesis in maize genotypes with different sensitivities to chilling.
Planta
198:
365-370
[CrossRef]
Lappartient AG,
Touraine B
(1996)
Demand-driven control of root ATP sulfurylase activity and SO42 uptake in intact canola.
Plant Physiol
111:
147-157
[Abstract]
Leegood RC
(1985)
The intercellular compartmentation of metabolites in leaves of Zea mays L.
Planta
164:
163-171
[CrossRef]
Li J,
Schiff JA
(1991)
Purification and properties of adenosine 5-phosphosulfate sulphotransferase from Euglena.
Biochem J
274:
355-360
Low PS,
Merida JR
(1996)
The oxidative burst in plant defense: function and signal transduction.
Physiol Plant
96:
533-542
[CrossRef]
MacKinney G
(1941)
Absorption of light by chlorophyll solutions.
Biol Chem
140:
312-322
Martin F,
Winspear MJ,
MacFarlane JD,
Oaks A
(1983)
Effect of methionine sulfoximine on the accumulation of ammonia in C3 and C4 leaves.
Plant Physiol
71:
177-181
[Abstract/Free Full Text]
Mills WR,
Joy KW
(1980)
A method for isolation of purified physiologically active chloroplasts, used to study the intracellular distribution of amino acids in pea leaves.
Planta
148:
75-83
[CrossRef]
Moore RC,
Black CC
(1979)
Nitrogen assimilation pathways in leaf mesophyll and bundle sheath cells of C4 photosynthesis plants formulated from comparative studies with Digitaria sanguinalis (L.) Scop.
Plant Physiol
64:
309-313
[Abstract/Free Full Text]
Newton GL,
Dorian R,
Fahey RC
(1981)
Analysis of biological thiols: derivatization with monobromobimane and separation by reverse-phase high-performance liquid chromatography.
Anal Biochem
114:
383-387
[CrossRef][ISI][Medline]
Neyra CA,
Hagemann RH
(1975)
Nitrate uptake and induction of nitrate reductase in excised corn roots.
Plant Physiol
56:
692-695
[Abstract/Free Full Text]
Nussbaum S,
Schmutz D,
Brunold C
(1988)
Regulation of assimilatory sulfate reduction by cadmium in Zea mays L.
Plant Physiol
88:
1407-1410
[Abstract/Free Full Text]
Ohnishi J,
Kanai R
(1983)
Differentiation of photorespiratory activity between mesophyll and bundle sheath cells of C4 plants. 1. Glycine oxidation by mitochondria.
Plant Cell Physiol
24:
1411-1420
[Abstract/Free Full Text]
Passera C,
Ghisi R
(1982)
ATP sulphurylase and O-acetylserine sulphydrylase in isolated mesophyll protoplasts and bundle sheath strands of S-deprived maize leaves.
J Exp Bot
33:
432-438
[Abstract/Free Full Text]
Pfündel E,
Nagel E,
Meister A
(1996)
Analyzing the light energy distribution in the photosynthetic apparatus of C4 plants using highly purified mesophyll and bundle-sheath thylakoids.
Plant Physiol
112:
1055-1070
[Abstract]
Rennenberg H,
Brunold C
(1994)
Significance of glutathione metabolism in plants under stress.
Prog Bot
55:
144-156
Rennenberg H,
Lamoureux GL
(1990)
Physiological processes that modulate the concentration of glutathione in plant cells.
In
H Rennenberg,
C Brunold,
LJ De Kok,
I Stulen,
eds, Sulphur Nutrition and Sulphur Assimilation in Higher Plants.
SPB Academic Publishing, The Hague, The Netherlands, pp 53-65
Rüegsegger A,
Brunold C
(1992)
Effect of cadmium on glutamylcysteine synthesis in maize seedlings.
Plant Physiol
99:
428-433
[Abstract/Free Full Text]
Schmutz D,
Brunold C
(1982)
Rapid and simple measurement of ATP sulfurylase activity in crude plant extracts using an ATP meter for bioluminescence determination.
Anal Biochem
121:
151-155
[CrossRef][ISI][Medline]
Schmutz D,
Brunold C
(1984)
Intercellular localization of assimilatory sulfate reduction in leaves of Zea mays and Triticum aestivum.
Plant Physiol
74:
866-870
[Abstract/Free Full Text]
Schmutz D,
Brunold C
(1985)
Localization of nitrite and sulfite reductase in bundle sheath and mesophyll cells of maize leaves.
Physiol Plant
64:
523-528
[CrossRef]
Schupp R,
Rennenberg H
(1988)
Diurnal changes in the glutathione content of spruce needles (Picea abies L.).
Plant Sci
57:
113-117
[CrossRef]
Setya A,
Murillo M,
Leustek T
(1996)
Sulfate reduction in higher plants: molecular evidence for a novel 5-adenylylsulfate reductase.
Proc Natl Acad Sci USA
93:
13383-13388
[Abstract/Free Full Text]
Sheen JY,
Bogorad L
(1988)
Differential expression in bundle sheath and mesophyll cells of maize of genes for photosystem II components encoded by the plastid genome.
Plant Physiol
86:
1020-1026
[Abstract/Free Full Text]
Valle EM,
Craig S,
Hatch MD,
Heldt HW
(1989)
Permeability and ultrastructure of bundle sheath cells isolated from C4 plants: structure-function studies and the role of plasmodesmata.
Bot Acta
102:
276-282
Valle EM,
Heldt HW
(1991)
Alanine synthesis by bundle sheath cells of maize.
Plant Physiol
95:
839-845
[Abstract/Free Full Text]
Vaughn KC,
Campbell WH
(1988)
Immunogold localization of nitrate reductase in maize leaves.
Plant Physiol
88:
1354-1357
[Abstract/Free Full Text]
Wyss HR,
Brunold C
(1979)
Regulation of adenosine 5-phosphosulfate sulfotransferase activity by H2S and cysteine in primary leaves of Phaseolus vulgaris L.
Planta
147:
37-42
[CrossRef]
Yamaya T,
Oaks A
(1988)
Distribution of two isoforms of glutamine synthetase in bundle sheath and mesophyll cells of corn leaves.
Physiol Plant
72:
23-28
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Abstract
Introduction