PDC activity and acetyl group accumulation in skeletal muscle during prolonged exercise.

Seven subjects cycled to exhaustion [58 +/- 7 (SE) min] at approximately 75% of their maximal oxygen uptake (VO2max). Needle biopsy samples were taken from the quadriceps femoris muscle at rest, after 3, 10, and 40 min of exercise, at exhaustion, and after 10 min of recovery. After 3 min of exercise, a nearly complete transformation of the pyruvate dehydrogenase complex (PDC) into active form had occurred and was maintained throughout the exercise period. The total in vitro activated PDC was unchanged during exercise. The muscle concentration of acetyl-CoA increased from a resting value of 8.4 +/- 1.0 to 31.6 +/- 3.3 mumol/kg dry wt at exhaustion and that of acetylcarnitine from 2.9 +/- 0.7 to 15.6 +/- 1.6 mmol/kg dry wt. This was accompanied by corresponding decreases in reduced CoA (CoASH) from 45.3 +/- 3.1 to 25.9 +/- 3.1 mumol/kg dry wt and in free carnitine from 18.8 +/- 0.7 to 5.7 +/- 0.5 mmol/kg dry wt. Acetyl group accumulation, in the form of acetyl-CoA and acetylcarnitine, was maintained throughout exercise to exhaustion while the glycogen content decreased by 90%. This suggests that availability of acetyl groups was not limiting to exercise performance despite the nearly total depletion of the glycogen store. The increased acetyl-CoA-to-CoASH ratio during exercise caused inhibition of neither the PDC transformation nor the calculated catalytic activity of active PDC.     

An acetyl group deficit limits mitochondrial ATP production at the onset of exercise

The oxygen deficit at the onset of submaximal exercise represents a period when the energy demand of contraction cannot be met solely by mitochondrial ATP generation, and as a consequence there is an acceleration of ATP re-synthesis from oxygen-independent routes (phosphocreatine hydrolysis and glycolysis). Historically, the origin of the oxygen deficit has been attributed to a lag in muscle blood flow and oxygen availability at the onset of exercise which limits mitochondrial respiration. However, more recent evidence suggests that considerable inertia exists at the level of mitochondrial enzyme activation and substrate supply. In support of this latter hypothesis, we have reported on a number of occasions that pharmacological activation of the pyruvate dehydrogenase complex (and consequent stockpiling of acetyl groups), using dichloroacetate or exercise interventions, can markedly reduce the degree of ATP re-synthesis from oxygen-independent routes during the rest-to-work transition period. This review will focus on these findings, and will offer the hypothesis that acetyl group delivery to the tricarboxylic acid cycle limits mitochondrial flux at the onset of exercise--the so-called acetyl group deficit.     

Association between muscle acetyl-CoA and acetylcarnitine levels in the exercising horse

Treadmill exercise of 2-min duration and increasing intensity resulted in increased formation of acetyl-CoA and acetylcarnitine in working muscle of Thoroughbred horses. At high work intensities a plateau was reached for both acetyl-CoA (approximately 50 mumols/kg dry muscle) and acetylcarnitine (approximately 20 mmol/kg dry muscle). Postexercise concentrations were significantly (P less than 0.001) correlated; [acetylcarnitine] = 349.[acetyl-CoA] + 2.4. The results indicate that approximately 350 mumols acetylcarnitine were accumulated for every 1 mumol acetyl-CoA. Under the conditions of exercise used it is probable that most of the acetyl-CoA formed is generated through the intramitochondrial decarboxylation of pyruvate. The acetyl groups of acetyl-CoA are apparently redistributed throughout the whole cell through formation of acetylcarnitine, which readily transverses the mitochondrial membrane. Despite the redistribution, however, the close correlation between acetylcarnitine and acetyl-CoA would indicate that equilibrium was maintained and that neither acetylcarnitine transferase nor carnitine/acetylcarnitine translocase were rate limiting. There is some question as to whether the changes observed relate directly to exercise itself or to the state in muscle 10 s or more after exercise.     

Malonyl-CoA and carnitine in regulation of fat oxidation in human skeletal muscle during exercise

Intracellular mechanisms regulating fat oxidation were investigated in human skeletal muscle during exercise. Eight young, healthy, moderately trained men performed bicycle exercise (60 min, 65% peak O2 consumption) on two occasions, where they ingested either 1) a high-carbohydrate diet (H-CHO) or 2) a low-carbohydrate diet (L-CHO) before exercise to alter muscle glycogen content as well as to induce, respectively, low and high rates of fat oxidation. Leg fat oxidation was 122% higher during exercise in L-CHO than in H-CHO (P < 0.001). In keeping with this, the activity of alpha2-AMP-activated protein kinase (alpha2-AMPK) was increased twice as much in L-CHO as in H-CHO (P < 0.01) at 60 min of exercise. However, acetyl-CoA carboxylase (ACC)beta Ser221 phosphorylation was increased to the same extent (6-fold) under the two conditions. The concentration of malonyl-CoA was reduced 13% by exercise in both conditions (P < 0.05). Pyruvate dehydrogenase activity was higher during exercise in H-CHO than in L-CHO (P < 0.01). In H-CHO only, the concentrations of acetyl-CoA and acetylcarnitine were increased (P < 0.001), and the concentration of free carnitine was decreased (P < 0.01), by exercise. The data suggest that a decrease in the concentration of malonyl-CoA, secondary to alpha2-AMPK activation and ACC inhibition (by phosphorylation), contributes to the increase in fat oxidation observed at the onset of exercise regardless of muscle glycogen levels. They also suggest that, with high muscle glycogen, the availability of free carnitine may limit fat oxidation during exercise, due to its increased use for acetylcarnitine formation.     

Acetylcarnitine formation during intense muscular contraction in humans

To study the changes in carnitine during intense muscular effort subjects underwent 4 min intermittent electrical stimulation of the quadriceps femoris muscle and on a separate occasion performed 4 min exercise on a bicycle ergometer. Biopsies of the vastus lateralis muscle were taken at rest and after 2 and 4 min of stimulation or exercise. Resting mean muscle total carnitine content was 20.0 mmol/kg dry muscle. Approximately 77% was free carnitine and 19% acetylcarnitine. Four minutes of stimulation or intense exercise did not effect total carnitine but did result in a marked fall in free carnitine and almost equivalent rise in acetylcarnitine. The results indicate that acetylcarnitine is a major metabolite formed during intense muscular effort and that carnitine may function in the regulation of the acetyl-CoA/CoA ratio by buffering excess production of acetyl units.     

Pyruvate dehydrogenase complex and acetyl-CoA carboxylase in pea root plastids: their characterization and role in modulating glycolytic carbon flow to fatty acid biosynthesis

The pyruvate dehydrogenase complex (PDC) and acetyl-CoA carboxylase(ACC, EC 6.4.1.2 [EC] ) have been characterized in pea root plastids.PDC activity was optimum in the presence of 1.0 mM pyruvate,1.5 mM NAD⁺ 0.1 mM CoA, 0.1 mM TPP, 5 mM MgCl₂, 3.0 mM cysteine-HCl,and 0.1 M Tricine (pH 8.0) and represents approximately 47%of the total cellular activity. ACC activity was greatest inthe presence of 1.0 mM acetyl-CoA, 4 mM NaHCO₃ mM ATP, 10 mMMgCl₂, 2.5 mM dithiothreitol, and 100 mM Tricine (pH 8.0). Bothenzymes were stimulated by reduced sulphydryl reagents and inhibitedby sulphydryl inhibitors. ACC was also inhibited by malonyl-CoAwhile PDC was inhibited by both malonyl-CoA and NADH. Both enzymeswere stimulated by DHAP and UDP-galactose while ACC was alsostimulated by PEP and F₁,6P. Palmitic acid and oleic acid bothinhibited ACC, but had essentially no effect on PDC. Palmitoyl-CoAinhibited both enzymes while PA and Lyso-PA inhibited PDC, butstimulated ACC. The results presented support the hypothesisthat PDC and ACC function in a co-ordinated fashion to promoteglycolytic carbon flow to fatty acid biosynthesis in pea rootplastids. Key words: Pisum sativum L., pyruvate dehydrogenase complex, acetyl-CoA carboxylase, roots, non-photosynthetic plastids     

Synthesis of citrate from phosphoenolpyruvate and acetylcarnitine by mitochondria from rabbit, pigeon and rat liver: Implications for lipogenesis

Rabbit, pigeon and rat liver mitochondria convert exogenous phosphoenolpyruvate and acetylcarnitine to citrate at rates of 14, 74 and 8 nmol/15 min/mg protein. Citrate formation is dependent on exogenous HCO3, is increased consistently by exogenous nucleotides (GDP, IDP, GTP, ADP, ATP) and inhibited strongly by 3-mercaptopicolinate and 1,2,3-benzenetricar☐ylate. Citrate is not made from pyruvate alone or combined with acetylcarnitine. Pigeon and rat liver mitochondria make large amounts of citrate from exogenous succinate, suggesting the presence of an endogenous source of acetyl units or a means of converting oxalacetate to acetyl units. Citrate synthesis from succinate by pigeon and rabbit mitochondria is increased significantly by exogenous acetylcarnitine. Pigeon and rat liver contain 80 and 15 times, respectively, more ATP:citrate lyase activity than does rabbit liver. Data suggest that mitochondrial phosphoenolpyruvate car☐ykinasein vivo could convert glycolysis-derived phosphoenolpyruvate to oxalacetate that, with acetyl CoA, could form citrate for export to support cytosolic lipogenesis as an activator of acetyl CoA car☐ylase, a carbon source via ATP:citrate lyase and NADPH via NADP: malate dehydrogenase or NADP: isocitrate dehydrogenase.     

AMPK activation is not critical in the regulation of muscle FA uptake and oxidation during low-intensity muscle contraction

To determine the role of AMP-activated protein kinase (AMPK) activation on the regulation of fatty acid (FA) uptake and oxidation, we perfused rat hindquarters with 6 mM glucose, 10 microU/ml insulin, 550 microM palmitate, and [14C]palmitate during rest (R) or electrical stimulation (ES), inducing low-intensity (0.1 Hz) muscle contraction either with or without 2 mM 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR). AICAR treatment significantly increased glucose and FA uptake during R (P < 0.05) but had no effect on either variable during ES (P > 0.05). AICAR treatment significantly increased total FA oxidation (P < 0.05) during both R (0.38 +/- 0.11 vs. 0.89 +/- 0.1 nmol x min(-1) x g(-1)) and ES (0.73 +/- 0.11 vs. 2.01 +/- 0.1 nmol x min(-1) x g(-1)), which was paralleled in both conditions by a significant increase and significant decrease in AMPK and acetyl-CoA carboxylase (ACC) activity, respectively (P < 0.05). Low-intensity muscle contraction increased glucose uptake, FA uptake, and total FA oxidation (P < 0.05) despite no change in AMPK (950.5 +/- 35.9 vs. 1,067.7 +/- 58.8 nmol x min(-1) x g(-1)) or ACC (51.2 +/- 6.7 vs. 55.7 +/- 2.0 nmol x min(-1) x g(-1)) activity from R to ES (P > 0.05). When contraction and AICAR treatment were combined, the AICAR-induced increase in AMPK activity (34%) did not account for the synergistic increase in FA oxidation (175%) observed under similar conditions. These results suggest that while AMPK-dependent mechanisms may regulate FA uptake and FA oxidation at rest, AMPK-independent mechanisms predominate during low-intensity muscle contraction.     

Formation of acetylcarnitine in muscle of horse during high intensity exercise

To study the changes in carnitine in muscle with spring exercise, two Thoroughbred horses performed two treadmill exercise tests. Biopsies of the middle gluteal were taken before, after exercise and after 12 min recovery. Resting mean muscle total carnitine content was 29.5 mmol.kg-1 dry muscle (d.m.). Approximately 88% was free carnitine, 7% acetylcarnitine and acylcarnitine was estimated at 5%. Exercise did not affect total carnitine, but resulted in a marked fall in free carnitine and almost equivalent rise in acetylcarnitine. The results are consistent with a role for carnitine in the regulation of the acetyl-CoA/CoA ratio during sprint exercise in the Thoroughbred horse by buffering excess production of acetyl units.     

Skeletal muscle metabolism during high-intensity sprint exercise is unaffected by dichloroacetate or acetate infusion.

This study investigated whether increased provision of oxidative substrate would reduce the reliance on nonoxidative ATP production and/or increase power output during maximal sprint exercise. The provision of oxidative substrate was increased at the onset of exercise by the infusion of acetate (AC; increased resting acetylcarnitine) or dichloroacetate [DCA; increased acetylcarnitine and greater activation of pyruvate dehydrogeanse (PDH-a)]. Subjects performed 10 s of maximal cycling on an isokinetic ergometer on three occasions after either DCA, AC, or saline (Con) infusion. Resting PDH-a with DCA was increased significantly over AC and Con trials (3.58 +/- 0.4 vs. 0.52 +/- 0.1 and 0.74 +/- 0.1 mmol. kg wet muscle(-1). min(-1)). DCA and AC significantly increased resting acetyl-CoA (35.2 +/- 4.4 and 22.7 +/- 2.9 vs. 10.2 +/- 1.3 micromol/kg dry muscle) and acetylcarnitine (12.9 +/- 1.4 and 11.0 +/- 1.0 vs. 3.3 +/- 0.6 mmol/kg dry muscle) over Con. Resting contents of phosphocreatine, lactate, ATP, and glycolytic intermediates were not different among trials. Average power output and total work done were not different among the three 10-s sprint trials. Postexercise, PDH-a in AC and Con trials had increased significantly but was still significantly lower than in DCA trial. Acetyl-CoA did not increase in any trial, whereas acetylcarnitine increased significantly only in DCA. Exercise caused identical decreases in ATP and phosphocreatine and identical increases in lactate, pyruvate, and glycolytic intermediates in all trials. These data suggest that there is an inability to utilize extra oxidative substrate (from either stored acetylcarnitine or increased PDH-a) during exercise at this intensity, possibly because of O(2) and/or metabolic limitations.     

Effect of ribose supplementation on resynthesis of adenine nucleotides after intense intermittent training in humans

The effect of oral ribose supplementation on the resynthesis of adenine nucleotides and performance after 1 wk of intense intermittent exercise was examined. Eight subjects performed a random double-blind crossover design. The subjects performed cycle training consisting of 15 x 10 s of all-out sprinting twice per day for 7 days. After training the subjects received either ribose (200 mg/kg body wt; Rib) or placebo (Pla) three times per day for 3 days. An exercise test was performed at 72 h after the last training session. Immediately after the last training session, muscle ATP was lowered (P < 0.05) by 25 +/- 2 and 22 +/- 3% in Pla and Rib, respectively. In both Pla and Rib, muscle ATP levels at 5 and 24 h after the exercise were still lower (P < 0.05) than pretraining. After 72 h, muscle ATP was similar (P > 0.05) to pretraining in Rib (24.6 +/- 0.6 vs. 26.2 +/- 0.2 mmol/kg dry wt) but still lower (P < 0.05) in Pla (21.1 +/- 0.5 vs. 26.0 +/- 0.2 mmol/kg dry wt) and higher (P < 0.05) in Rib than in Pla. Plasma hypoxanthine levels after the test performed at 72 h were higher (P < 0.05) in Rib compared with Pla. Mean and peak power outputs during the test performed at 72 h were similar (P > 0.05) in Pla and Rib. The results support the hypothesis that the availability of ribose in the muscle is a limiting factor for the rate of resynthesis of ATP. Furthermore, the reduction in muscle ATP observed after intense training does not appear to be limiting for high-intensity exercise performance.     

Acetate Availability and Utilization Supports the Growth of Mutant Sub-Populations on Aging Bacterial Colonies

When bacterial colonies age most cells enter a stationary phase, but sub-populations of mutant bacteria can continue to grow and accumulate. These sub-populations include bacteria with mutations in rpoB (RNA polymerase β-subunit) or rpoS (RNA polymerase stress-response sigma factor). Here we have identified acetate as a nutrient present in the aging colonies that is utilized by these mutant subpopulations to support their continued growth. Proteome analysis of aging colonies showed that several proteins involved in acetate conversion and utilization were upregulated during aging. Acetate is known to be excreted during the exponential growth phase but can be imported later during the transition to stationary phase and converted to acetyl-CoA. Acetyl-CoA is used in multiple processes, including feeding into the TCA cycle, generating ATP via the glyoxylate shunt, as a source of acetyl groups for protein modification, and to support fatty acid biosynthesis. We showed that deletion of acs (encodes acetyl-CoA synthetase; converts acetate into acetyl-CoA) significantly reduced the accumulation of rpoB and rpoS mutant subpopulations on aging colonies. Measurement of radioactive acetate uptake showed that the rate of conversion decreased in aging wild-type colonies, was maintained at a constant level in the rpoB mutant, and significantly increased in the aging rpoS mutant. Finally, we showed that the growth of subpopulations on aging colonies was greatly enhanced if the aging colony itself was unable to utilize acetate, leaving more acetate available for mutant subpopulations to use. Accordingly, the data show that the accumulation of subpopulations of rpoB and rpoS mutants on aging colonies is supported by the availability in the aging colony of acetate, and by the ability of the subpopulation cells to convert the acetate to acetyl-CoA.     

Skeletal muscle metabolism is unaffected by DCA infusion and hyperoxia after onset of intense aerobic exercise

This study investigated whether hyperoxic breathing (100% O(2)) or increasing oxidative substrate supply [dichloroacetate (DCA) infusion] would increase oxidative phosphorylation and reduce the reliance on substrate phosphorylation at the onset of high-intensity aerobic exercise. Eight male subjects cycled at 90% maximal O(2) uptake (VO(2 max)) for 90 s in three randomized conditions: 1) normoxic breathing and saline infusion over 1 h immediately before exercise (CON), 2) normoxic breathing and saline infusion with DCA (100 mg/kg body wt), and 3) hyperoxic breathing for 20 min at rest and during exercise and saline infusion (HYP). Muscle biopsies from the vastus lateralis were sampled at rest and after 30 and 90 s of exercise. DCA infusion increased pyruvate dehydrogenase (PDH) activation above CON and HYP (3.10 +/- 0.23, 0.56 +/- 0.08, 0.69 +/- 0.05 mmol x kg wet muscle(-1) x min(-1), respectively) and significantly increased both acetyl-CoA and acetylcarnitine (11.0 +/- 0.7, 2.0 +/- 0.5, 2.2 +/- 0.5 mmol/kg dry muscle, respectively) at rest. However, DCA and HYP did not alter phosphocreatine degradation and lactate accumulation and, therefore, the reliance on substrate phosphorylation during 30 s (CON, 51.2 +/- 5.4; DCA, 56.5 +/- 7.1; HYP, 69.5 +/- 6.3 mmol ATP/kg dry muscle) and 90 s of exercise (CON, 90.6 +/- 9.5; DCA, 107.2 +/- 13.0; HYP, 101.2 +/- 15.2 mmol ATP/kg dry muscle). These data suggest that the rate of oxidative phosphorylation at the onset of exercise at 90% VO(2 max) is not limited by oxygen availability to the active muscle or by substrate availability (metabolic inertia) at the level of PDH in aerobically trained subjects.     

Progressive increase in human skeletal muscle AMPKalpha2 activity and ACC phosphorylation during exercise

The effect of prolonged moderate-intensity exercise on human skeletal muscle AMP-activated protein kinase (AMPK)alpha1 and -alpha2 activity and acetyl-CoA carboxylase (ACCbeta) and neuronal nitric oxide synthase (nNOSmu) phosphorylation was investigated. Seven active healthy individuals cycled for 30 min at a workload requiring 62.8 +/- 1.3% of peak O(2) consumption (VO(2 peak)) with muscle biopsies obtained from the vastus lateralis at rest and at 5 and 30 min of exercise. AMPKalpha1 activity was not altered by exercise; however, AMPKalpha2 activity was significantly (P < 0.05) elevated after 5 min (approximately 2-fold), and further elevated (P < 0.05) after 30 min (approximately 3-fold) of exercise. ACCbeta phosphorylation was increased (P < 0.05) after 5 min (approximately 18-fold compared with rest) and increased (P < 0.05) further after 30 min of exercise (approximately 36-fold compared with rest). Increases in AMPKalpha2 activity were significantly correlated with both increases in ACCbeta phosphorylation and reductions in muscle glycogen content. Fat oxidation tended (P = 0.058) to increase progressively during exercise. Muscle creatine phosphate was lower (P < 0.05), and muscle creatine, calculated free AMP, and free AMP-to-ATP ratio were higher (P < 0.05) at both 5 and 30 min of exercise compared with those at rest. At 30 min of exercise, the values of these metabolites were not significantly different from those at 5 min of exercise. Phosphorylation of nNOSmu was variable, and despite the mean doubling with exercise, statistically significance was not achieved (P = 0.304). Western blots indicated that AMPKapproximately 2 was associated with both nNOSmu and ACCbeta consistent with them both being substrates of AMPKalpha2 in vivo. In conclusion, AMPKalpha2 activity and ACCbeta phosphorylation increase progressively during moderate exercise at approximately 60% of VO(2 peak) in humans, with these responses more closely coupled to muscle glycogen content than muscle AMP/ATP ratio.     

Purification and characterization of pyruvate dehydrogenase complex from borccoli floral buds.

The pyruvate dehydrogenase complex (PDC) was purified from Brassica oleracea var. italica floral buds to a specific activity of approximately 6 μmol of NADH formed/min/ mg of protein. The PDC had cofactor requirements for NAD⁺, thiamine pyrophosphate, coenzyme A, and a divalent cation (Mg²⁺, Ca²⁺, or Mn²⁺). The enzyme catalyzed the oxidative decarboxylation of pyruvate at a rate threefold faster than 2-oxobutyrate but was inactive toward 2-oxoglutarate. The PDC was competively inhibited by acetyl-CoA against CoA and NADH against NAD⁺. The enzyme was shown to be more sensitive to regulation by NADH than acetyl-CoA.     

Multiple Mass Isotopomer Tracing of Acetyl-CoA Metabolism in Langendorff-perfused Rat Hearts: CHANNELING OF ACETYL-CoA FROM PYRUVATE DEHYDROGENASE TO CARNITINE ACETYLTRANSFERASE*

We developed an isotopic technique to assess mitochondrial acetyl-CoA turnover (≈citric acid flux) in perfused rat hearts. Hearts are perfused with buffer containing tracer [¹³C₂,²H₃]acetate, which forms M5 + M4 + M3 acetyl-CoA. The buffer may also contain one or two labeled substrates, which generate M2 acetyl-CoA (e.g. [¹³C₆]glucose or [1,2-¹³C₂]palmitate) or/and M1 acetyl-CoA (e.g. [1-¹³C]octanoate). The total acetyl-CoA turnover and the contributions of fuels to acetyl-CoA are calculated from the uptake of the acetate tracer and the mass isotopomer distribution of acetyl-CoA. The method was applied to measurements of acetyl-CoA turnover under different conditions (glucose ± palmitate ± insulin ± dichloroacetate). The data revealed (i) substrate cycling between glycogen and glucose-6-P and between glucose-6-P and triose phosphates, (ii) the release of small excess acetyl groups as acetylcarnitine and ketone bodies, and (iii) the channeling of mitochondrial acetyl-CoA from pyruvate dehydrogenase to carnitine acetyltransferase. Because of this channeling, the labeling of acetylcarnitine and ketone bodies released by the heart are not proxies of the labeling of mitochondrial acetyl-CoA.     

Contributions of Carnitine Acetyltransferases to Intracellular Acetyl Unit Transport in Candida albicans

Transport of acetyl-CoA between intracellular compartments is mediated by carnitine acetyltransferases (Cats) that reversibly link acetyl units to the carrier molecule carnitine. The genome of the opportunistic pathogenic yeast Candida albicans encodes several (putative) Cats: the peroxisomal and mitochondrial Cat2 isoenzymes encoded by a single gene and the carnitine acetyltransferase homologs Yat1 and Yat2. To determine the contributions of the individual Cats, various carnitine acetyltransferase mutant strains were constructed and subjected to phenotypic and biochemical analyses on different carbon sources. We show that mitochondrial Cat2 is required for the intramitochondrial conversion of acetylcarnitine to acetyl-CoA, which is essential for a functional tricarboxylic acid cycle during growth on oleate, acetate, ethanol, and citrate. Yat1 is cytosolic and contributes to acetyl-CoA transport from the cytosol during growth on ethanol or acetate, but its activity is not required for growth on oleate. Yat2 is also cytosolic, but we were unable to attribute any function to this enzyme. Surprisingly, peroxisomal Cat2 is essential neither for export of acetyl units during growth on oleate nor for the import of acetyl units during growth on acetate or ethanol. Oxidation of fatty acids still takes place in the absence of peroxisomal Cat2, but biomass formation is absent, and the strain displays a growth delay on acetate and ethanol that can be partially rescued by the addition of carnitine. Based on our results, we present a model for the intracellular flow of acetyl units under various growth conditions and the roles of each of the Cats in this process.