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Plant Physiol. (1998) 118: 1439-1445
Export of Carbon from Chloroplasts at Night1
Jürgen Schleucher2,
Peter J. Vanderveer, and
Thomas D. Sharkey*
Department of Botany (J.S., P.J.V., T.D.S.), and Department of
Biochemistry (J.S.), University of Wisconsin, Madison, Wisconsin
53706
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ABSTRACT |
Hexose
export from chloroplasts at night has been inferred in previous studies
of mutant and transgenic plants. We have tested whether hexose export
is the normal route of carbon export from chloroplasts at night. We
used nuclear magnetic resonance to distinguish glucose (Glc) made from
hexose export and Glc made from triose export. Glc synthesized in vitro
from fructose-6-phosphate in the presence of deuterium-labeled water
had deuterium incorporated at C-2, whereas synthesis from triose
phosphates caused C-2 through C-5 to become deuterated. In both tomato
(Lycopersicon esculentum L.) and bean (Phaseolus
vulgaris L.), Glc from sucrose made at night in the presence of
deuterium-enriched water was deuterated only in the C-2 position,
indicating that >75% of carbon is exported as hexoses at night. In
darkness the phosphate in the cytosol was 28 mM, whereas
that in the chloroplasts was 5 mM, but hexose phosphates
were 10-fold higher in the cytosol than in the chloroplasts. Therefore,
hexose phosphates would not move out of chloroplasts without the input
of energy. We conclude that most carbon leaves chloroplasts at night as
Glc, maltose, or higher maltodextrins under normal conditions.
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INTRODUCTION |
Chloroplasts provide all of the reduced carbon in higher plants:
from photosynthesis during the day and from starch degradation at
night. During the day reduced carbon is exported as triose phosphate,
especially dihydroxyacetone phosphate (Walker and Herold, 1977 ), which
is used to make Suc and similar transport sugars. A membrane-bound
antiporter that exchanges triose phosphate for phosphate has been
isolated and sequenced, and the gene has been cloned (Flügge and
Heldt, 1991). The triose-phosphate translocator is unregulated; the
flow of carbon is determined by the metabolism in the chloroplast and
cytosol. During the day, production of triose phosphates inside the
chloroplast and production of phosphate outside result in concentration
gradients that drive triose phosphate export and phosphate import.
Starch breakdown can be amylolytic, resulting in maltodextrins,
maltose, and Glc, or phosphorolytic, resulting in hexose and triose
phosphates (Levi and Gibbs, 1976; Preiss, 1982). The phosphorolytic pathway has a high capacity when phosphate levels are high (Heldt et
al., 1977) and preserves energy contained in the Glc-Glc bond in
starch. Phosphorolytic starch breakdown is presumed to lead to the
production of triose phosphates, which can be exported on the phosphate
translocator. Amylolytic starch breakdown leads to maltose and Glc,
both of which can be transported, presumably on the Glc transporter
(Schäfer et al., 1977; Herold et al., 1981). A number of studies
have concluded that the phosphorolytic pathway is the primary pathway
for starch breakdown (Heldt et al., 1977; Levi and Preiss, 1978; Stitt
and Heldt, 1981a).
Recent work with mutant and transgenic plants has indicated that when
triose phosphate export or metabolism in the cytosol is blocked, carbon
export during the day declines and export at night increases. For
example, (a) a mutant of Flaveria linearis deficient in
cytosolic FBPase activity but with no obvious phenotypic changes was
found to export little carbon during the day (Sharkey et al., 1992;
Zrenner et al., 1996); (b) antisense triose-phosphate- translocator
plants were found to have reduced carbon export during the day and
enhanced export during the night (Riesmeier et al., 1993; Heineke et
al., 1994a; Häusler et al., 1998); and (c) a mutant of
Arabidopsis that accumulates starch was found to lack a hexose
transporter (Trethewey and ap Rees, 1994).
These observations indicate that at night there is a path for carbon
export from chloroplasts that does not function during the day and that
does not involve either the triose-phosphate translocator or cytosolic
FBPase. The nighttime pathway could work by export of Glc, with or
without esterified phosphate. However, known hexose-phosphate
transporters in plastids have low activity (Batz et al., 1992) and are
generally considered important primarily in starch accumulation (Heldt
et al., 1991; Overlach et al., 1993; Schott et al., 1995; Häusler
et al., 1998). The Glc/maltose transporter described by Schäfer
et al. (1977) and Herold et al. (1981) could explain the nighttime
export.
We have developed a method to test whether plants normally export
hexoses or trioses from chloroplasts during starch breakdown, and have
made measurements to distinguish between phosphorylated and
unphosphorylated compounds. During ketose phosphate/aldose phosphate
isomerization by sugar isomerases, such as triose phosphate isomerase
and phosphoglucoisomerase, one hydrogen of C-1 of a ketose and C-2 of
the corresponding aldose can exchange with hydrogen in the medium
(Hanson and Rose, 1975). If sugar leaves the chloroplast as hexose and
is then converted to Suc following isomerization by
phosphoglucoisomerase, i.e. if hexose units stay intact from starch
breakdown to Suc synthesis, the Glc moiety of Suc should exchange only
the hydrogen on C-2 with the medium. However, if triose phosphates were
involved, the hydrogen on C-2 would exchange as before and triose
phosphate isomerase would allow exchange of hydrogens on what will
become C-3 and C-5 of Glc. In addition, aldolase would allow for the
exchange of hydrogen at C-4. Therefore, the Glc moiety of Suc made in
the dark in 2H-labeled medium would be enriched
in 2H only at C-2 if carbon export were only by
hexoses, but at carbons 2 through 5 if triose phosphate were involved.
We tested whether chloroplasts normally export hexoses or trioses at
night by allowing starch degradation in the presence of water labeled
with 2H and using NMR to determine the
intramolecular distribution of H on Glc
liberated from starch. We also measured the concentration of Glc-6-P
and Fru-6-P in chloroplasts and the cytosol in leaves harvested in
darkness to determine whether there is a sufficient gradient in these
phosphorolytic breakdown products to cause substantial export.
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MATERIALS AND METHODS |
Plant Material
Bean (Phaseolus vulgaris L. cv Linden) and tomato
(Lycopersicon esculentum L. cv UC82B) plants were grown in
growth chambers with a 16-h day. The day temperature was 22°C and the
night temperature was 18°C. Light was provided by a mixture of
high-pressure sodium and metal halide lamps and was 500 µmol
m 2 s1. The RH was 60%.
Plants were grown in 3-L pots in Metro-Mix 360 (Scotts- Sierra
Horticultural Products, Marysville, OH).
Experimental Protocol
Two types of labeling experiments were carried out. In the first
type, leaves were surrounded with a plastic bag containing deuterated
water. By diffusion, the water in the leaf became labeled. Starch
breakdown was allowed to proceed normally in darkness for 2.5 or 6 h. The leaf was then cut off the plant, frozen, and stored at  80°C
until analyzed. In the second type of experiment, leaves were detached
and the petiole was put into 2H-enriched water
containing EDTA to enhance phloem export from the cut tip. The leaf was
allowed to export Suc for 24 h. The irrigation water containing
the leaf exudate was then frozen and stored for later analysis.
Derivatization for NMR
Per gram of crude sugar, 5 g of anhydrous ZnCl2,
0.5 mL of 85% H3PO4, and
200 mL of anhydrous acetone were added and the reaction mixture was
stirred for 2 d at room temperature under exclusion of moisture
(Glen et al., 1951). After the 1st d, any solid material was loosened
from the wall of the flask with a spatula. This procedure resulted in
the formation of an equilibrium mixture of approximately 7%
1,2-O-isopropyliden- -D-glucofuranose
(compound 2, Fig. 1), 86%
1,2-5,6-di-O-isopropylidene--D-glucofuranose
(compound 1, Fig. 1), and 7% Glc (measured by integration of the
signals of the anomeric protons by proton NMR). The reaction mixture
was poured into 1.2 volumes of ice water and the pH was adjusted
to 2.1 with 6 mol L1 HCl. This solution was
stirred at room temperature to hydrolyze compound 1 to compound 2. After 30 h, the reaction mixture contained approximately 88%
compound 2, 5% compound 1, and 7% Glc. The diisopropylidene derivative of Fru that was formed from Suc was stable under these conditions. The solution was neutralized by pouring it into a solution
of Na2CO3 (160 g
L1).
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| Figure 1.
Isolation of Glc derivatives for
2H-NMR spectroscopy. Reaction conditions and yields are
given in ``Materials and Methods''. Compound 1, 1,2-5,6-di-O-isopropyliden--D-glucofuranose;
compound 2, 1,2-O-isopropylidene--D-glucofuranose.
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The precipitate of zinc compounds was removed by filtration, the
solution was evaporated to dryness under reduced pressure, and the
residue (sodium salts and compound 2) was dried in a vacuum at 60°C
for 30 min. The powdered residue was extracted four times with boiling
ethyl acetate (30 mL g1 sugar educt), and the
hot solution was filtered through a glass funnel into a flask
containing a small amount of solid NaHCO3. The
ethyl acetate extract was concentrated and compound 2 was allowed to
crystallize from the hot solution. Compound 2 was isolated by
filtration, washed with cold diethyl ether, and dried. An additional crystallization from ethyl acetate was often required to remove the
diisopropylidene derivative of Fru.
NMR Methodology
For the NMR measurements, compound 2 was dissolved at 50°C in
2.4 mL of acetonitrile (HPLC grade) in a 10-mm NMR tube (catalog no.
Z18, 398-9, Aldrich) containing a few crystals of
NaHCO3, H-depleted
water (50-250 µL for amounts of compound 2 from 100-300 mg), and 20 µL of a 2% (v/v) solution of hexadeuterobenzene in acetonitrile.
This sample composition and the measurement temperature of 50°C
reduced the viscosity of the solution and therefore the H signal width. The addition of water increased
the solubility of compound 2 and ensured that a single signal outside
of the range of the carbon-bound hydrogens was observed for
2HHO and the hydroxy protons of compound 2. Longitudinal
relaxation times of the 2H signals were
determined using an inversion-recovery pulse sequence (Vold et al.,
1968). The longest longitudinal relaxation times were 300 to 400 ms.
2H-NMR spectra were acquired at 50°C on DMX-400
and DMX-500 spectrometers (Bruker Instruments, Billerica, MA) in an
unlocked mode using a 90° excitation pulse. A repetition time (and
acquisition time) of 1 s (approximately 2 longitudinal relaxation
times of the slowest relaxing signal) was used. Proton decoupling using
the globally optimized alternating-phase rectangular pulses (Shaka et
al., 1985) sequence was applied during the acquisition of the
2H spectra to obtain singlet Lorentzian signals.
It has been reported (Martin and Martin, 1990) and was confirmed here
(not shown) that this did not cause measurable intensity changes as a
result of nuclear Overhauser enhancements. The free induction decays
(raw data) were zero-filled and Fourier-transformed with a line
broadening of 1 Hz using the program Felix (Molecular Simulations, San
Diego, CA). The signals of the solvent and the added hexadeuterobenzene were used for phasing. The sample sizes given in ``Results'' refer to
independently prepared samples from separate plants. The assignment of
the compound 2 signals were taken from Mackie and Perlin (1965) and
were confirmed by analysis of the proton NMR spectrum. The
stereospecific assignment of the C-6 methylene hydrogens of compound 2 was obtained from a 2H-NMR spectrum of a
stereospecifically deuterated sample (see below).
Control Experiments
Analysis for hydrogen exchange with the solvent was accomplished
by carrying out the isolation sequence in the presence of 2%
D2O at each step. 2H-NMR
spectra of compound 2 formed in the presence of
D2O and in H2O were
compared. To identify possible isotope effects of the derivatization
reactions, control experiments were performed in which some reaction
steps were carried out with low turnover so as to maximize the
2H discrimination caused by isotope effects.
Partial turnover of Suc was achieved by reacting Suc with acetone and
sulfuric acid in the presence of water. Partial turnover of compound 1 to 2 was achieved by stopping the hydrolysis at low turnover.
2H-NMR spectra of the control samples were
compared with spectra obtained from normal preparations (yields  88%). Epimerization of the C-6 methylene hydrogens was ruled out by
synthesis of stereospecifically deuterated Glc.
(6S)-1,6-Anhydro-2,3,4-tri-O-benzoyl- -D-glucopyranose-6-2H
was synthesized according to the method of Ohrui et al. (1983) and was
hydrolyzed to
(6S)-1,6-anhydro--D-glucopyranose-6-2H.
This was reacted directly to yield
(6S)-1,2-O-isopropyliden--D-glucofuranose-6-2H.Com-parison
of the 2H-NMR spectra of this compound and of
the precursor
(6S)-1,6-anhydro-2,3,4-tri-O-benzoyl--D-glucopyranose-6-2H
showed that no epimerization at C-6 had occurred.
We confirmed the predicted intramolecular labeling for hexose phosphate
isomerase and triose phosphate isomerase by derivatizing Glc made from
Fru-6-P or from triose phosphates in the presence of
2H-labeled water. The intramolecular labeling was
determined by NMR analysis.
Subcellular Metabolite Levels
Subcellular metabolite levels were determined by nonaqueous
fractionation of bean leaves using the method of Gerhardt and Heldt
(1984), as modified by Sharkey and Vanderveer (1989). Freeze-dried leaves were sonicated in cold heptane and then layered on a
heptane/tetrachloroethylene density gradient. After centrifugation, the
gradient was separated into six fractions. Each fraction was assayed
for NADP-glyceraldehyde-3-P dehydrogenase (chloroplast marker), PEP
carboxylase (cytosol marker), and -mannosidase (vacuole marker) plus
each metabolite of interest. No indication of hexose phosphate in the
vacuole was seen and therefore the data were evaluated assuming only
two compartments, the chloroplast and the cytosol, for hexose
phosphates. Glc and Fru were not resolved well on the gradient,
probably because a large fraction of each was located in the vacuole
(Heineke et al., 1994b); therefore, only whole-leaf levels of these
compounds are reported. The phosphate nutrition of the bean plants was
restricted to lower the phosphate level in the vacuole (Sharkey and
Vanderveer, 1989) but there was still a substantial amount present. To
determine the amounts of phosphate in the chloroplast, cytosol, and
vacuole, an empirical three-compartment analysis was used (Riens et
al., 1991). In this analysis each possible distribution of phosphate among the three compartments was considered and the predicted distribution in the gradient was compared with the observed
distribution. The assumed subcellular distribution that predicted a
distribution through the density gradient closest to the observed one
was taken as the best estimate of the subcellular distribution.
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RESULTS |
The labeling pattern resulting from the action of
phosphoglucoisomerase in vitro is shown in the first trace of Figure
2. In this spectrum, C-2 is heavily
deuterated, whereas the other hydrogens have natural abundance levels
of 2H. Whereas the labeling is only about 4-fold
over natural abundance, the exclusive labeling of C-2 is obvious.
Enzyme isotope effects can only deplete 2H in the
C-2 position (J. Schleucher, unpublished data), so accumulation of
2H at C-2 must have resulted from incorporation
from the medium. The second trace shows that there was substantial
deuteration of C-2 through C-5 in Glc made from triose phosphate in
vitro when 2H-labeled water was present in the
medium.
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| Figure 2.
2H-NMR spectra of
2H-labeled samples of the Glc derivative
1,2-O-isopropylidene-D-glucofuranose. Top
trace, Glc was synthesized from Fru-6-P via phosphoglucoisomerase and
Glc-6-phosphatase in 2H-labeled water. Middle trace, Glc
was synthesized in 2H-labeled water from dihydroxyacetone
phosphate via triose phosphate isomerase, aldolase, Fru bisphosphatase,
phosphoglucoisomerase, and Glc 6-phosphatase. Bottom trace, Intact
bean leaves were placed at the beginning of darkness in plastic bags
containing paper soaked with 2H-labeled water, the leaves
were harvested after 3 h and treated with invertase, and the Glc
liberated from Suc was derivatized.
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We tested whether starch conversion to Suc at night involved triose
phosphates by putting a bag with 2H-labeled water
around a bean leaf at night after the lights of the growth chamber had
been turned off. The 2H-labeled water was allowed
to diffuse into the leaf and we assumed that substantial
2H was present throughout the leaf sap. Leaves
were left with bags around them for 2.5 h, during which time the
amount of starch broken down was about the same as the Suc content in
the leaf at the end of the day. About 63% of the Suc in the leaf after the incubation should have originated from starch breakdown.
Preexisting Suc made in the light would have had no enrichment in
2H, since 2H was present
only after the lights were off. The third trace in Figure 2 shows that
the labeling pattern of the Glc moiety of Suc was essentially identical
to the labeling pattern caused by phosphoglucoisomerase. In other
experiments with bean and tomato plants, leaves were harvested at other
times during the night and similar results were found (Table
I).
To determine whether sufficient gradients exist to cause export of
hexose phosphates at night, we measured the concentrations of hexose
phosphates and phosphate in the chloroplast and cytosol. Bean leaves
were quickly frozen in the dark and then subjected to nonaqueous
fractionation. The gradient for hexose phosphates was in the wrong
direction for simple diffusion of these compounds (Table
II). The gradient for phosphate was large
and in the proper direction to cause exchange of phosphate from the
cytosol for hexose phosphate from the stroma. Assuming 50 µL liquid
volume mg1 chlorophyll for the stromal volume
and 25 µL mg1 chlorophyll for the cytosolic
volume (average of values reported for barley and spinach [Winter et
al., 1993, 1994]), we could calculate that phosphate was 28 mM in the cytosol and 5 mM in the stroma. The
hexose phosphates were 1 mM in the cytosol and not in
isomerase equilibrium. In the stroma, Glc-6-P was 0.1 mM and Fru was undetectable.
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Table II.
Metabolite concentrations in the chloroplast and
cytosol
Bean leaves were analyzed by nonaqueous fractionation. Leaves were
quickly frozen in the dark.
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Fru levels were low and constant day and night, whereas Glc levels were
higher during the day than at night (Fig.
3). Suc was very high during the day and
lower but still substantial during the night. The Glc level varied
between day and night but much less than the Suc level. Because most of
the hexoses are located in the vacuole, it was not possible to resolve
the intracellular location of the free hexoses.
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| Figure 3.
Levels of Glc, Fru, and Suc in bean leaves during
the day and at night. Dark bars, Data determined using leaf samples
taken at midnight; lighter bars, data from samples taken at noon.
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DISCUSSION |
The NMR data clearly indicate that most carbon leaves chloroplasts
at night as hexoses, not as trioses. Previously,
13C-NMR was used to show that amyloplasts take up
hexoses and not trioses (Keeling et al., 1988). The hexose exported
from chloroplasts at night cannot be phosphorylated, since the
concentration gradient for both Glc-6-P and Fru-6-P is in the wrong
direction. A large gradient in phosphate could theoretically drive
hexose phosphate export against its concentration gradient. However,
the phosphate concentration was 6-fold higher in the cytosol (Table II)
but needs to be 10-fold higher to force hexose phosphate export from the chloroplast. The level of Glc could be sufficient to cause hexose
export; however, accurate measurements of the distribution between the
chloroplast and cytosol are difficult because of the amount of Glc in
the vacuole. Some plastids can transport hexose phosphates but the gene
for the Glc-6-P/P antiporter of plastids is not expressed in leaves
and, therefore, this transporter is not present in chloroplasts
(Kammerer et al., 1998). Thus, all available evidence indicates that
free hexoses, possibly including maltose and higher maltodextrins, are
the primary transport compounds during Suc synthesis from starch.
Hexose export has been postulated to explain the results from mutant
and transgenic plants (Sharkey et al., 1992; Heineke et al., 1994a;
Zrenner et al., 1996); we now show that hexose export is the normal
route of carbon export from chloroplasts at night in untransformed,
nonmutant plants. From the data in Figure 1 and Table I, we calculate
that at least 75%, and possibly 100%, of the carbon is exported from
chloroplasts at night as hexose, not triose. The finding that plants
lacking the Glc transporter accumulate starch (Trethewey and ap Rees,
1994) indicates that the phosphorolytic pathway of starch breakdown and
export cannot replace the amylolytic pathway.
Cytosolic FBPase and Suc-phosphate synthase are important regulatory
points in the synthesis of Suc (Stitt et al., 1987). Suc phosphate
synthase activity can decline at night but often does not (Huber et
al., 1989). However, cytosolic FBPase is nearly always regulated to
have a very low activity in darkness. This enzyme is strongly inhibited
by Fru 2,6-bisP (Stitt, 1990), which is high at night in most plants
studied (Sicher et al., 1986; Sicher and Bunce, 1987). Thus, cytosolic
FBPase activity is reduced much more at night than is Suc phosphate
synthase. If hexose is exported at night, then Suc phosphate synthase,
not cytosolic FBPase, activity is needed. Therefore, the regulation of
the two important regulatory steps in Suc synthesis is consistent with hexose export as the primary path for carbon export from chloroplasts at night. These pathways are shown in Figure
4. Given the low activity of cytosolic
FBPase restricting triose phosphate supply for Suc synthesis and the
unfavorable concentration gradient and lack of carrier for hexose
phosphates, it is easy to see why the phosphorolytic pathway cannot
compensate for the loss of the Glc transporter in the Arabidopsis
mutant TC265 (Trethewey and ap Rees, 1994).
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| Figure 4.
Pathways of carbon metabolism described in
``Discussion''. A triose phosphate/phosphate antiporter is the source
of carbon for Suc synthesis during the day, whereas the Glc/maltose
uniporter is the source of carbon for Suc synthesis during the night.
DHAP, Dihydroxyacetone-3-P; F6P, Fru-6-P; FBP, Fru-1,6-bisP; GAP,
glyceraldehyde-3-P; G6P, Glc-6-P; PGA, phosphoglyceric acid; RuBP,
ribulose 1,5-bisP; and UDPG, UDP-Glc.
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The pathway of starch degradation is not yet certain. Most studies have
found that the chloroplastic -amylase (endoamylase) is the only
enzyme to attack intact starch granules. The maltodextrins released by
-amylase are debranched by the chloroplastic R-enzyme and broken
down by -amylase to maltose and higher maltodextrins (Okita and
Preiss, 1980). Whereas most starch-degrading enzymes occur both inside
and outside of the chloroplast, maltase activity occurs only outside of
the chloroplast (Okita et al., 1979; Kakefuda et al., 1986). Thus,
maltose could be an important component of the carbon export from
chloroplasts at night.
The energetics of the phosphorolytic versus amylolytic pathways have
been discussed by Beck (1985). The phosphorolytic pathway leading to
triose phosphate export would require three A(U)TPs per Suc molecule
(two ATPs at phosphofructokinase inside the chloroplast plus one UTP at
UDP-Glc pyrophosphylase; Fig. 4). The amylolytic pathway with Glc
exported would require the same amount of energy (two ATP at hexokinase
in the cytosol plus one UTP). If maltose were exported and broken down
by phosphorolysis in the cytosol, the total energy required could be
reduced by up to one-third. Thus, the amylolytic pathway is potentially
more energy efficient than the phosphorolytic pathway of starch
breakdown. Phosphate speeds starch breakdown, but the Glc-1-P formed
may be used to synthesize maltose for export (Kruger and ap Rees,
1983).
Starch breakdown has been reported in illuminated leaves. If, as seems
likely, this indicates amylolytic breakdown followed by export of
carbon from the chloroplast, then this carbon would bypass the
cytosolic FBPase. The coordinate regulation of the cytosolic and
chloroplastic FBPases is crucial in determining what proportion of
carbon is exported from chloroplasts during the day (Stitt et al.,
1987). Daytime starch degradation and export of amylolytic products
must ordinarily be severely restricted, since mutant and transgenic
plants lacking parts of the triose phosphate export pathway export most
or all of their carbon at night. One signal that influences starch
breakdown is the concentration of phosphate. In the stroma the
concentration of phosphate is higher at night than during the day
(Santarius and Heber, 1965; Sharkey and Vanderveer, 1989). The lower
phosphate level during the day could inhibit starch breakdown (Stitt
and Heldt, 1981b); however, this would affect phosphorolytic starch
breakdown, whereas data reported here indicate that most starch
breakdown is amylolytic.
The amylolytic breakdown of starch could be restricted during the day
by the high pH of the chloroplast, since the amylases have relatively
low pH optima. Pongratz and Beck (1978) reported that amylases in
chloroplasts exhibit a diurnal oscillation, being more active at night.
This does not seem to be the result of thiol reduction processes, since
amylase activity is not affected by DTT (Okita and Preiss, 1980).
Extending this finding, Kakefuda and Preiss (1997) reported evidence of
an endoamylase that changes activity by more than 5-fold between day
and night (i.e. is more active at night). The regulation of this enzyme
could explain how plants avoid bypassing the regulation by cytosolic
FBPase during the day.
In conclusion, the results presented here indicate that carbon from
starch breakdown at night is normally exported from chloroplasts as
hexose (Fig. 4). The amylolytic pathway is the dominant pathway for
starch breakdown and export despite the high levels of phosphate in
chloroplasts at night because phosphorylated compounds are not
exported. Amylolytic breakdown of starch is regulated so that very little occurs during the day under normal conditions,
forcing carbon through the cytosolic FBPase.
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FOOTNOTES |
1
This research was supported by the
U.S. Department of Energy (grant no. DE-FG-02-87ER13785).
J.S. was supported in part by a grant from the Deutsche
Forschungsgemeinschaft.
2
Present address: Department of Medical
Biochemistry and Biophysics, Umeå University, S-90187 Umeå, Sweden.
*
Corresponding author; e-mail tsharkey{at}facstaff.wisc.edu; fax
1-608-262-7509.
Received May 22, 1998;
accepted September 4, 1998.
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ABBREVIATIONS |
Abbreviation:
FBPase, Fru-1,6-bisphosphatase.
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ACKNOWLEDGMENT |
We would like to thank Prof. John Markley for the use of the
National Magnetic Resonance Facility at Madison, which is supported by
the National Institutes of Health, the University of Wisconsin, the
National Science Foundation, and the U.S. Department of Agriculture.
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Abstract
Introduction