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1 Department of Biochemistry and 2 Department of Nutrition, University of Montreal, Montreal, Québec, Canada H3C 3J7; 3 Department of Nutrition, Case Western Reserve University, Cleveland, Ohio 44106-7139; and 4 Department of Physiology, George Washington University, Medical Center, Washington, DC 20037-2337
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ABSTRACT |
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 TOP
 ABSTRACT
 INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
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Little is known about the sources of cytosolic acetyl-CoA used for the synthesis of malonyl-CoA, a key regulator of fatty acid oxidation in the heart. We tested the hypothesis that citrate provides acetyl-CoA for malonyl-CoA synthesis after its mitochondrial efflux and cleavage by cytosolic ATP-citrate lyase. We expanded on a previous study where we characterized citrate release from perfused rat hearts (Vincent G, Comte B, Poirier M, and Des Rosiers C. Citrate release by perfused rat hearts: a window on mitochondrial cataplerosis. Am J Physiol Endocrinol Metab 278: E846-E856, 2000). In the present study, we show that citrate release rates, ranging from 6 to 22 nmol/min, can support a net increase in malonyl-CoA concentrations induced by changes in substrate supply, at most 0.7 nmol/min. In experiments with [U-13C](lactate + pyruvate) and [1-13C]oleate, we show that the acetyl moiety of malonyl-CoA is derived from both pyruvate and long-chain fatty acids. This 13C-labeling of malonyl-CoA occurred without any changes in its concentration. Hydroxycitrate, an inhibitor of ATP-citrate lyase, prevents increases in malonyl-CoA concentrations and decreases its labeling from [U-13C](lactate + pyruvate). Our data support at least a partial role of citrate in the transfer from the mitochondria to cytosol of acetyl units for malonyl-CoA synthesis. In addition, they provide a dynamic picture of malonyl-CoA metabolism: even when the malonyl-CoA concentration remains constant, there appears to be a constant need to supply acetyl-CoA from various carbon sources, both carbohydrates and lipids, for malonyl-CoA synthesis.
gas chromatography-mass spectrometry; adenosine 5'-triphosphate-citrate lyase; hydroxycitrate; acetyl-coenzyme A; citric acid cycle; 13C-substrate; isotopomer analysis
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INTRODUCTION |
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TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
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THE INTRACELLULAR
CONCENTRATION of malonyl-CoA modulates a number of physiological
and pathophysiological events. These effects of malonyl-CoA are related
to its inhibition of carnitine palmitoyl transferase 1. The latter
controls the entry of long-chain acyl-CoA into mitochondria, where they
are 
-oxidized for energy production. In addition, carnitine
palmitoyl transferase 1 activity affects cytosolic concentrations of
long-chain acyl-CoAs, which influence signal transduction and binding
of nuclear transcription factors (5, 16, 19, 42). In
lipogenic tissues such as the liver, malonyl-CoA is both a modulator of
fatty acid oxidation and an intermediate of fatty acid synthesis
(25). In nonlipogenic tissues such as the heart and
skeletal muscle, cytosolic malonyl-CoA modulates fatty acid oxidation
and is a component of a fuel-sensing and signaling mechanism that
responds to changes in the cell's substrate supply and energy
expenditure (31, 33). Recently, a dysregulated malonyl-CoA
metabolism has been implicated in insulin resistance (33),
apoptosis (19), and functional recovery of the
heart after ischemia (20).
In the heart, the regulatory role of cytosolic malonyl-CoA is supported
by the kinetic and regulatory properties of enzymes involved in its
metabolism (13, 14, 20, 23, 33, 39). Malonyl-CoA is
synthesized by the predominant 280-kDa isoform of acetyl-CoA
carboxylase (ACC) or ACC
. This enzyme is regulated at
the transcriptional and posttranslational levels, the latter via
phosphorylation by 5'-AMP-activated protein kinase and protein kinase
A. It is also activated by citrate and inhibited by long-chain acyl-CoA. Because tissue acetyl-CoA levels are lower than the Km of ACC for acetyl-CoA (117 µM), the ACC activity seems to be substrate driven.
The only known fate of cardiac malonyl-CoA is recycling to acetyl-CoA
via malonyl-CoA decarboxylase (MCD). Much less is known about the
regulation of MCD compared with ACC. Small variations in
the energy demand of the heart could modulate MCD activity and results
in rapid changes in malonyl-CoA levels (13, 17). It is
unclear as to whether cardiac MCD is regulated by phosphorylation as in
the extensor digitorum longus muscle (14, 35).
Several issues regarding the proposed mechanism of fuel regulation by
malonyl-CoA still remain unresolved. First, how can one explain that
long-chain fatty acids (LCFA) are -oxidized at an intracellular
malonyl-CoA concentration that should inhibit completely the prevailing
muscle isoform of carnitine palmitoyl transferase 1? Proposed
explanations include malonyl-CoA protein binding and/or
compartmentation (3, 13, 15, 18, 25). Second, how are
cytosolic acetyl-CoA and malonyl-CoA levels correlated with fat and
carbohydrate oxidation and the citric acid cycle (CAC) flux in
mitochondria? Third, what is the source(s) of cytosolic acetyl-CoA for malonyl-CoA synthesis? One hypothesis is that acetyl-CoA generated by mitochondrial pyruvate dehydrogenase is channeled to
carnitine acetyl transferase and translocase (2, 23, 24). A second hypothesis is that mitochondrial acetyl-CoA is transferred to
the cytosol via citrate, the citrate transporter and cytosolic ATP-citrate lyase. The activities of enzymes involved in mitochondrial transfer of acetyl units via citrate or acetylcarnitine are small compared with the CAC flux rate (1, 4, 10, 27, 36). However, these activities could be sufficient to maintain the very
small pool of malonyl-CoA (between 0.2 and 4 nmol/g wet wt) (4,
17, 20, 23, 37).
We previously demonstrated a modulation of mitochondrial citrate efflux in perfused rat hearts, reflected by rates of citrate release (40). In the present study, we examined the link between mitochondrial citrate synthesis/efflux and malonyl-CoA synthesis. To test the transport of acetyl units via citrate, we used hydroxycitrate, an inhibitor of ATP-citrate lyase (41). We also investigated the sources of the acetyl moiety of malonyl-CoA with [13C]pyruvate and oleate by mass isotopomer analysis (8) to test the hypothesis of a preferential channeling of pyruvate-derived acetyl-CoA to malonyl-CoA.
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EXPERIMENTAL PROCEDURES |
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TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
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Chemicals
The sources of chemicals, biological products, and 13C-substrates have been identified previously (11, 12, 40). [3H]acetyl-CoA (25 µCi/mol) was obtained from ICN (Montreal, Quebec, Canada). Hydroxycitrate (Citrin K; Garcinia Cambogia Extract Potassium Salt) was a generous gift from Dr. V. Badmaev of Sabinsa (Piscataway, NJ). The dialyzed albumin solution (13.4%, BSA fraction V, fatty acid poor, Bayer; Kankakee, IL) and the stock solution of 20 mM sodium oleate complexed to albumin were prepared and stored as described previously (40).Heart Perfusions
Animal experimentation was approved by the local animal ethics committee in compliance with guidelines of the Canadian Council on Animal Care. Procedures for isolation and perfusion of rat hearts in the Langendorff mode have been described elsewhere (11, 12, 40). Briefly, the hearts of fed male Sprague-Dawley rats (180-210 g, Charles River Breeding Laboratories; St-Constant, Quebec, Canada) were perfused at a constant pressure of 70 mmHg with nonrecirculating modified Krebs-Henseleit buffer (pH 7.4) containing 119 mM NaCl, 4.8 mM KCl, 1.3 mM CaCl2, 1.2 mM KH2PO4, 1.2 mM MgSO4, 25 mM NaHCO3, 5.5 mM glucose, and 8 nM insulin as well as other additives indicated below. The setup for continuous monitoring of functional parameters, using instruments linked to a microcomputer, has been described earlier (40). At the end of the experiments, the hearts were freeze clamped and stored in liquid nitrogen.Perfusion Protocols
A two 30-min step protocol was designed to document the effect of selected metabolic perturbations that differentially affected the citrate release rate (40) on malonyl-CoA 1) net synthesis, as reflected by an increase in its tissue levels over 30 min; and 2) 13C-labeling. All hearts underwent an initial 30-min perfusion with 5.5 mM glucose, 8 nM insulin, 0.5 mM lactate, and 0.05 mM pyruvate to ascertain that the tissue level of malonyl-CoA would be the same at the beginning of all metabolic interventions. One group of hearts was freeze clamped after 30 min. Parameters measured in effluent or tissue for this 0- to 30-min period served as baseline values. All other groups of hearts underwent an additional 30 min of perfusion before freeze clamping. In addition to the substrates listed in Table 1, the perfusion buffer contained 5 mM glucose and 8 nM insulin. The buffer composition for protocol 1, which served as a control, is that used for the baseline period. Buffer modifications for the 30- to 60-min period included an increase in the concentration of pyruvate from 0.05 to 0.5 mM, the addition of a fatty acid, either 0.4 mM oleate, a LCFA complexed to 4% albumin, or 0.2 mM octanoate, a medium-chain fatty acid, and the addition of 1 mM hydroxycitrate. Note that albumin was present only in the LCFA group. The ionized calcium and endogenous free fatty acid concentrations of the 4% albumin-containing buffer were determined to be 1.2 and 0.3 mM, respectively. The total free fatty acid concentration for the LCFA group was 0.7 mM. For protocols 1A, 3A, and 5A, the buffer composition was identical to that of protocols 1, 3, and 5, respectively, except that unlabeled lactate, pyruvate, and oleate were replaced with [U-13C3](lactate + pyruvate) and [1-13C]oleate. For protocol 3A, [U-13C3](lactate + pyruvate) was present for the 0- to 60-min period and [1-13C]oleate was present for the 30- to 60-min period. For protocols 1A and 5A, in which hearts were perfused in the absence or in the presence of 1 mM hydroxycitrate, respectively, [U-13C3](lactate + pyruvate) was present for the last 20 min of the 30- to 60-min time period. Thus for protocol 5A, hearts were perfused for 10 min in the presence of 1 mM hydroxycitrate to allow for ATP-citrate lyase inhibition before the addition of 13C-substrates.
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The following constraints were linked to the use of hydroxycitrate. First, hydroxycitrate was available as the potassium salt. The total potassium concentration in the perfusion buffer was maintained at 4.8 mM by lowering its potassium chloride content to 2.1 mM. Second, because hydroxycitrate is a calcium chelator (9), its addition to the perfusion buffer lowered the ionized calcium concentration from 0.94 ± 0.01 to 0.68 ± 0.01 mM (P < 0.05, n = 9, unpaired t-test), although it did not change the total calcium concentration (1.23 ± 0.03 mM, n = 9). Increasing the total calcium concentration of the buffer containing hydroxycitrate to 1.7 mM reestablished its ionized calcium concentration to 0.98 ± 0.02 mM (n = 5). Third, there was a background contamination by citrate in the hydroxycitrate solution (0.5% or 50 µM in a 1 mM solution) that prevented the assessment of citrate release rates.
Analytic Procedures
Gas chromotography-mass spectrometry (GC-MS) methods for the determination of citrate release (40) and 13C mass isotopomer distribution (MID) of tissue malonyl-CoA (32) have been described previously. Tissue malonyl-CoA levels were measured radioenzymatically using a modification of the procedure of McGarry and Brown (26). In brief, 0.05 g wet wt of powdered hearts was homogenized in 1 ml of 10% trichloroacetic acid, followed by centrifugation at 14,000 rpm for 15 min at 4°C. Pellets were kept for protein determination by the Bradford method (6). Supernatants were washed six times with 1 ml of ether, dried in a Speed-Vac, and stored at
80°C. The
incubation mixture for malonyl-CoA assay (final volume 1 ml) contained
50 µl lyophilized extract [resuspended in 1 M potassium
phosphate buffer-10 mM EDTA (pH 7.1)], 200 µmol potassium phosphate
buffer (pH 7.1), 2 µmol EDTA, 2.5 µmol dithiothreitol, 0.2 µmol
NADPH, 1 mg BSA, 0.75 nmol [3H]acetyl-CoA (0.9 µCi/nmol), and 5 mU rat liver fatty acid synthase (21).
All assays were calibrated by spiking duplicate samples with 0.0625 nmol malonyl-CoA. We verified the linearity of the assay with tissue
weights in the range of 0.05-0.1 g wet wt. Analyses of total and
ionized calcium levels in perfusion buffer samples were conducted by
the Clinical Biochemistry Laboratory of Notre-Dame Hospital.
Calculations, Data Presentation, and Statistical Analyses
13C-labeling of metabolites.
Mass isotopomers of malonyl-CoA containing 1 and 2 13C
atoms are identified as Mi with i = 1 and 2. The absolute molar percent enrichment (MPE) of a given mass
isotopomer Mi was calculated as follows
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(1) |
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RESULTS |
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ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
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Cardiac Contractile Parameters
In the baseline period, perfused hearts maintained a constant spontaneous beating at 309 ± 5 beats/min, a coronary flow rate of 11 ± 1 ml/min, a left ventricular developed pressure of 113 ± 2 mmHg, a rate-pressure product of (34.9 ± 0.8) × 10
3 mmHg · beats · min1
(not shown), and a dP/dtmax of 3,267 ± 167 mmHg/s (Fig. 1, left). Compared with the baseline period (0-30 min), the contractile activity (dP/dtmax) decreased in the 30- to
60-min period in all groups except in hearts perfused with 0.5 mM
pyruvate (Fig. 1, middle). Hydroxycitrate addition to
control hearts resulted in a 20% decline of
dP/dtmax values (Fig. 1, right). This
effect of hydroxycitrate was not due to calcium chelation
(9), because it was also observed after normalization of
the buffer concentration of ionized calcium (not shown). Changes in
dP/dtmax values paralleled those of the left
ventricular developed pressure and of the rate-pressure product,
whereas values of coronary flow rate and heart rate remained similar to
the baseline values (not shown).
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Citrate Efflux and Malonyl-CoA Concentrations
Under all conditions, citrate release rates were constant for a given 30-min period, ranging from 7 to 22 nmol/min. In control hearts, citrate release rates (Fig. 2A) and tissue levels (Fig. 2B) were the same in the 30- to 60-min period than in the baseline period. Increasing pyruvate concentration from 0.05 to 0.5 mM, or adding octanoate or oleate to the perfusion buffer during the 30- to 60-min period, increased citrate release rates and tissue citrate levels in parallel. A similar correlation between citrate release rates and tissue levels was observed in our previous study in normoxic Langendorff-perfused rat hearts (40), although not in the swine heart in vivo (28).
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In hearts perfused under control conditions and in those perfused with 0.5 mM pyruvate, malonyl-CoA levels were similarly increased by 30% at 60 min compared with 30 min (Fig. 2C), reflecting net malonyl-CoA synthesis. Malonyl-CoA levels were not increased in the presence of oleate but markedly increased in the presence of octanoate, in agreement with recent data from Longnus et al. (22).
Addition of hydroxycitrate did not modify tissue citrate levels (Fig.
3A). However, it prevented the
increase in tissue malonyl-CoA levels during the 30- to 60-min period
(Fig. 3B). This effect was not observed in perfusions with 1 mM citrate (not shown).
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Sources of Acetyl Moiety for Malonyl-CoA Synthesis
We recently developed a technique to assess the 13C MID of malonyl-CoA (32). With the use of this technique, we probed the origin of the acetyl moiety of malonyl-CoA using [U-13C3](lactate + pyruvate) and [1-13C]oleate in the same perfusion experiments (protocol 3A). Decarboxylation of [U-13C3]pyruvate forms M2 isotopomers of acetyl-CoA and malonyl-CoA, whereas [1-13C]oleate oxidation forms M1 isotopomers. Note that in these nonrecirculating perfusions with 25 mM bicarbonate buffer, the incorporation of 13CO2 derived from the oxidation of the 13C-substrates is negligible. This is demonstrated by the absence of the M3 isotopomer of malonyl-CoA.In hearts perfused with [U-13C3](lactate + pyruvate) and [1-13C]oleate, malonyl-CoA was enriched
in both M1 and M2 isotopomers (Fig.
4A). From these M2 and M1
enrichment values, one can estimate the percentage of malonyl-CoA
molecules that arose from acetyl-CoA generated by pyruvate
decarboxylation and oleate oxidation, respectively, following a
reasoning similar to that used previously for estimating the relative
contribution of these pathways to the acetyl moiety of citrate (cf.
Eqs. 5 and 6 of Ref. 11,
respectively). Note that this calculation does not require any
corrections for recycling of [13C]citrate isotopomers in
the TCA cycle, but it is subject to some uncertainties because it
assumes the existence of only one pool of malonyl-CoA (3,
13, 15, 18).
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From the M2 enrichment of malonyl-CoA (9.9%), one can calculate a 10% contribution of exogenous pyruvate decarboxylation to the acetyl moiety of malonyl-CoA. To calculate the contribution of exogenous fatty acid oxidation, one has to consider two factors: 1) only one of nine acetyl units of the oleate molecule was labeled with 13C, and 2) the albumin preparation contained 0.3 mM unlabeled free fatty acids, which must be added to the 0.4 mM concentration of [1-13C]oleate. Therefore, the contribution of exogenous fatty acids to the acetyl moiety of malonyl-CoA was obtained by multiplying the M1 enrichment of malonyl-CoA (5.8%) by [9 × (0.4 + 0.3)/0.4], yielding 91%. Note that this value of 91% probably overestimates slightly the true contribution of exogenous fatty acids to malonyl-CoA because some unlabeled free fatty acids in the albumin preparation could have less than nine acetyl units (e.g., palmitate).
We also tested the effect of hydroxycitrate on malonyl-CoA labeling from [U-13C3](lactate + pyruvate) (protocols 1A and 5A). Hydroxycitrate decreased slightly (22%), but significantly, the M2 enrichment of malonyl-CoA (Fig. 4B).
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DISCUSSION |
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TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
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The goal of the present study was to shed some light on the
sources of the acetyl moiety of cytosolic malonyl-CoA in the heart. There are two possible pathways for transferring acetyl units from
mitochondria to the cytosol: 1) citrate transporter and
ATP-citrate lyase and 2) carnitine acetyl translocase and
transferase (Fig. 5). The activities of
enzymes involved in the two putative processes, albeit low (1, 4,
10, 27, 36), seem sufficient to maintain the very small pool of
malonyl-CoA in the heart. We decided to investigate first the citrate
pathway because our previous study (40) had identified a
sizeable citrate efflux from the perfused rat heart (5-25
nmol/min). This efflux of citrate amounts to 0.2-1% of CAC flux
and is modulated by the nature of the substrates offered to the heart
and by energy demand.
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In the present study, we confirmed the relationship between citrate
release rates and tissue content (40) (Fig. 2,
A and B). In addition, we compared changes in
citrate release and in malonyl-CoA tissue levels induced by
modifications of substrate supply (Fig. 2, B and
C). No correlation was found, but citrate release rates were
far in excess of increases in malonyl-CoA content based on the
following. Malonyl-CoA concentrations were measured 30 min after
changing the substrate profile in the perfusate. Most likely, the
various substrate mixtures affected differentially the supply of
substrate (acetyl-CoA) and/or the levels of allosteric regulators of
ACC (citrate and long-chain acyl-CoA). Malonyl-CoA accumulation over time could have resulted from an increased substrate flux through ACC with no change or a decrease in flux
through MCD. We do not know the time course of these changes, but it is likely to have been fairly rapid (4, 13, 17, 23, 37). Because the half-life of malonyl-CoA in similarly perfused rat hearts
is ~1.5 min (32), let us assume that the changes in
malonyl-CoA concentrations occurred over 5 min. The net rate of
malonyl-CoA accumulation would then be 0.2-0.7
nmol · min1 · g wet
wt1. This compares to citrate release rates
ranging from 7 to 22 nmol/min. Note that citrate release rates
represent the difference between citrate efflux from mitochondria and
citrate disposal via cytosolic ATP-citrate lyase. These observations
are compatible with citrate being involved in the transfer of acetyl units.
We used hydroxycitrate, an inhibitor of ATP-citrate lyase
(41), to further test the participation of the citrate
pathway in the heart. Saha et al. (34) have shown a
decrease in malonyl-CoA concentrations in rat muscle incubated in the
presence of hydroxycitrate. In our study, the addition of 1 mM
hydroxycitrate to the perfusate prevented the increase in malonyl-CoA
concentration over baseline levels (Fig. 3B). This effect
was observed despite 1) a lowering of the contractile
activity of the heart, a condition that could potentially increase the
activity of ACC and/or decrease that of MCD (17,
33); and 2) the known capacity of hydroxycitrate to
act as an allosteric activator of ACC (34).
In fact, each of these two factors should have favored an increase,
rather than a decrease, in tissue malonyl-CoA levels. These
observations are also compatible with the participation of cytosolic
ATP-citrate lyase in the generation of acetyl-CoA for malonyl-CoA synthesis.
Additional arguments for the citrate pathway were obtained from
experiments with 13C-labeled substrates. First, in
perfusions with [U-13C3](lactate + pyruvate) and [1-13C]oleate, we showed that acetyl units
used for malonyl-CoA synthesis are generated from both decarboxylation
of exogenous pyruvate/lactate and -oxidation of exogenous LCFA. The
relative contributions of these two processes to malonyl-CoA formation
(10% and 90%) are similar to those reported for the acetyl moiety of
citrate in rat hearts perfused under similar conditions
(40). Despite some uncertainties in the calculation of
these estimates, these data argue against a preferential channeling of
pyruvate-derived acetyl-CoA to malonyl-CoA via acetylcarnitine
transferase/translocase (2, 23, 24). In studies conducted
by Lysiak et al. (24) in isolated rat heart mitochondria,
the formation of acetylcarnitine from pyruvate was favored by low
concentrations of catalytic intermediates of the CAC. Thus, in the
presence of high carnitine concentration, the formation of
acetylcarnitine had to be the major fate of acetyl-CoA derived from
pyruvate decarboxylation. Furthermore, the decrease in acetylcarnitine
formation upon addition of malate is probably explained by the
incorporation of acetyl groups into citrate. In the present study, the
mix of substrates provided (some of them anaplerotic) presumably
insured adequate constant concentrations of CAC intermediates as
reflected by a sizeable citrate efflux.
Hydroxycitrate prevented the increase in tissue malonyl-CoA levels and lowered the extent of 13C-labeling of malonyl-CoA from [U-13C3](lactate + pyruvate). Thus, despite the presence of 1 mM hydroxycitrate, some acetyl units derived from [U-13C3](lactate + pyruvate) were converted to malonyl-CoA. These data can be interpreted in two ways. First, the concentration of hydroxycitrate (1 mM) might not have been sufficient to fully inhibit ATP-citrate lyase. Hydroxycitrate, a tri-anion at physiological pH, does not permeate easily through cell membranes. This is why the concentration of malonyl-CoA in incubating muscle (34) decreased as hydroxycitrate concentrations were raised from 1 to 4 mM. This also explains why the Ki for hydroxycitrate of isolated ATP-citrate lyase (0.05-0.6 µM) (38, 41) is three to four orders of magnitude lower than the concentration of hydroxycitrate required to inhibit fatty acid synthesis by 50% in the intact liver (2 mM) (7). Second, it is possible that the citrate-cleavage pathway and the acetylcarnitine transferase/translocase pathway both contribute to the acetyl moiety of malonyl-CoA, at least under some circumstances.
In conclusion, our data support the role of citrate in the transfer of
acetyl units from the mitochondria to cytosol in the heart. Results
from this and our previous study (32) show that measurements of malonyl-CoA concentrations and related enzyme activities provide only a partial picture of malonyl-CoA metabolism. Cycling between acetyl-CoA and malonyl-CoA does not change the malonyl-CoA concentration in the heart, where the only known fate of
malonyl-CoA is degradation to acetyl-CoA. However, cytosolic acetyl-CoA
might be hydrolyzed to acetate as occurs in numerous organs
(30) or converted to acetylcarnitine before export. Thus, although the low level of malonyl-CoA in cytosol remains fairly constant, there is probably a constant need to supply acetyl-CoA to
ACC. This study with 13C-substrates provides
a dynamic picture of the carbon sources and fluxes leading to
malonyl-CoA even when its concentration does not change. Results from
this study do not exclude, however, a contribution of the
acetylcarnitine pathway in the transfer of acetyl units from the
mitochondria to cytosol. The citrate and the acetylcarnitine pathways
might coexist and ensure a constant supply of acetyl-CoA to
ACC under various conditions. One factor that could
determine the contributions of the two pathways could be the relative
availability of anaplerotic substrates versus acetyl-CoA and carnitine
in mitochondria. Another potential factor is the regulation of
ATP-citrate lyase activity by covalent modification (29).
Further studies will investigate the respective roles of citrate and
acetylcarnitine pathways to the supply of acetyl moiety of malonyl-CoA.
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ACKNOWLEDGEMENTS |
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The authors thank Dr. Marc Prentki and Laura Segall for assisting in setting up the malonyl-CoA assay and Drs. John Chatham, Blandine Comte, and Hans Kornberg for helpful comments.
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FOOTNOTES |
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The study was supported by the Canadian Institutes for Health Research Grants MT-9575 and MT-10920 (to C. Des Rosiers and a studentship to G. Vincent).
Part of this work was presented at the 1999 and 2001 meetings of Experimental Biology.
Address for reprint requests and other correspondence: C. Des Rosiers, Laboratoire du Métabolisme Intermédiaire, Y-3616, Centre Hospitalier de l'Université de Montréal, Hôpital Notre-Dame, 1560 rue Sherbrooke Est, Montréal, Québec, Canada H2L 4M1 (E-mail: christine.des.rosiers{at}umontreal.ca).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
June 13, 2002;10.1152/ajpheart.00244.2002
Received 25 February 2002; accepted in final form 10 June 2002.
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REFERENCES |
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TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
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0.05, all data at 30-60 min versus those at
0-30 min; #P < 0.05, effect of HC: values
compared with control for the 30- to 60-min period. MCFA and LCFA,
medium- and long-chain fatty acid, respectively.
