Triacylglycerol Breakdown Research Articles

Additivity of adrenaline and contractions on hormone-sensitive lipase, but not on glycogen phosphorylase, in rat muscle.

Hormone-sensitive lipase (HSL) has been proposed to regulate triacylglycerol (TG) breakdown in skeletal muscle. In muscles with different fibre type compositions the influence on HSL of two major stimuli causing TG mobilization was studied. Incubated soleus and extensor digitorum longus (EDL) muscles from 70 g rats were stimulated by adrenaline (5.5 microm, 6 min) or contractions (200 ms tetani, 1 Hz, 1 min) in maximally effective doses or by both adrenaline and contractions. Hormone-sensitive lipase activity was increased significantly by adrenaline as well as contractions, and the highest activity (P < 0.05) was seen with combined stimulation [Soleus: 0.40 +/- 0.03 (SE) m-unit mg protein(-1) (basal), 0.65 +/- 0.02 (adrenaline), 0.65 +/- 0.03 (contractions), 0.78 +/- 0.03 (adrenaline and contractions); EDL: 0.18 +/- 0.01, 0.30 +/- 0.02, 0.26 +/- 0.02, 0.32 +/- 0.01]. Glycogen phosphorylase activity was always increased more by adrenaline compared with contractions [Soleus: 60 +/- 4 (a/a + b)% vs. 46 +/- 3 (P < 0.05); EDL: 60 +/- 5 vs. 39 +/- 6 (P < 0.05)]. After combined stimulation glycogen phosphorylase activity in soleus [59 +/- 3 (a/a + b)%] was identical to and in EDL [45 +/- 4 (a/a + b)%] smaller (P < 0.05) than the activity after adrenaline only. In slow-twitch oxidative as well as in fast-twitch glycolytic muscle HSL is activated by both adrenaline and contractions. These stimuli are partially additive indicating at least partly different mechanisms of action. Contractions may impair the enhancing effect of adrenaline on glycogen phosphorylase activity in muscle.

Acute changes in triacylglycerol lipase activity of human adipose tissue during exercise

Although physical exercise is known to increase adipose tissue lipolysis, its effect on the activity of triacylglycerol (TG) lipase, the enzyme regulating TG breakdown, is not known. The aim of the present study was to monitor the acute changes in TG lipase activity of adipose tissue induced during moderate exercise. For this purpose a new assay, sensitive to the phosphorylation state of the enzyme, was developed. Ten young sedentary men cycled for 30 min at a heart rate of 120-130 beats min(-1). Needle adipose tissue biopsy was performed from the buttock area at rest, at 5, 15, and 30 min of exercise, as well as at 15 min of passive recovery. Five other men served as controls by being biopsied as above without exercising. TG lipase activity was determined by measuring the decrease of endogenous TG concentration during incubation of the homogenized tissue. TG lipase activity increased 6.4-fold above baseline at 5 min of exercise (P < 0.001) and fell gradually afterwards, whereas it did not change significantly in the control group. In conclusion, our data show that TG lipase activity in human adipose tissue peaks early during exercise and subsequently decreases despite the maintenance of the physical stimulus.

Requirement for 3-ketoacyl-CoA thiolase-2 in peroxisome development, fatty acid beta-oxidation and breakdown of triacylglycerol in lipid bodies of Arabidopsis seedlings.

3-ketoacyl-CoA thiolase (KAT) (EC: 2.3.1.16) catalyses a key step in fatty acid beta-oxidation. Expression of the Arabidopsis thaliana KAT gene on chromosome 2 (KAT2), which encodes a peroxisomal thiolase, is activated in early seedling growth. We identified a T-DNA insertion in this gene which abolishes its expression and eliminates most of the thiolase activity in seedlings. In the homozygous kat2 mutant, seedling growth is dependent upon exogenous sugar, and storage triacylglycerol (TAG) and lipid bodies persist in green cotyledons. The peroxisomes in cotyledons of kat2 seedlings are very large, the total peroxisomal compartment is dramatically increased, and some peroxisomes contain unusual membrane inclusions. The size and number of plastids and mitochondria are also modified. Long-chain (C16 to C20) fatty acyl-CoAs accumulate in kat2 seedlings, indicating that the mutant lacks long-chain thiolase activity. In addition, extracts from kat2 seedlings have significantly decreased activity with aceto-acetyl CoA, and KAT2 appears to be the only thiolase gene expressed at significant levels during germination and seedling growth, indicating that KAT2 has broad substrate specificity. The kat2 phenotype can be complemented by KAT2 or KAT5 cDNAs driven by the CaMV 35S promoter, showing that these enzymes are functionally equivalent, but that expression of the KAT5 gene in seedlings is too low for effective catabolism of TAG. By comparison with glyoxylate cycle mutants, it is concluded that while gluconeogenesis from fatty acids is not absolutely required to support Arabidopsis seedling growth, peroxisomal beta-oxidation is essential, which is in turn required for breakdown of TAG in lipid bodies.

Glycerol: a neglected variable in metabolic processes?

Glycerol is a small and simple molecule produced in the breakdown of glucose, proteins, pyruvate, triacylglycerols and other glycerolipid, as well as release from dietary fats. An increasing number of observations show that glycerol is probably involved in a surprising variety of physiopathologic mechanisms. Glycerol has long been known to play fundamental roles in several vital physiological processes, in prokaryotes and eukaryotes, and is an important intermediate of energy metabolism. Despite some differences in the details of their operation, many of these mechanisms have been preserved throughout evolution, demonstrating their fundamental importance. In particular, glycerol can control osmotic activity and crystal formation and then act as a cryoprotective agent. Furthermore, its properties make it useful in numerous industrial, therapeutic and diagnostic applications. Few studies have focussed directly on glycerol, however, and while its metabolism is increasingly well documented, much of the details remain unknown. Considering the importance of glycerol in multiple vital physiological processes, its study could help unlock important physiopathological mechanisms.

Determination of glycerol in plasma by an automated enzymatic spectrophotometric procedure.

The plasma concentration of glycerol, the backbone of triglycerides and the end product of triacylglycerol breakdown, is considered to reflect lipolysis in adipose tissue. We evaluated an automated enzymatic procedure for the measurement of glycerol in plasma. The assay was linear up to 250 micromol/l. The detection limit was 8 micromol/l. Recovery averaged 94% from spiked plasma and 104% from diluted plasma. An extensive precision study performed according to the guidelines of the National Committee for Clinical Laboratory Standards showed within-run coefficients of variation between 14.8% and 2.6% for concentrations ranging from 28 micromol/l to 164 micromol/l. The reference range for fasting healthy adult men was 14-69 micromol/l. Glycerol levels were significantly correlated with free fatty acid levels. This automated enzymatic method is rapid and reliable, and provides greater sensitivity or convenience than previously described procedures.

Development of leptin resistance in rat soleus muscle in response to high-fat diets.

Direct evidence for leptin resistance in peripheral tissues such as skeletal muscle does not exist. Therefore, we investigated the effects of different high-fat diets on lipid metabolism in isolated rat soleus muscle and specifically explored whether leptin's stimulatory effects on muscle lipid metabolism would be reduced after exposure to high-fat diets. Control (Cont, 12% kcal fat) and high-fat [60% kcal safflower oil (n-6) (HF-Saff); 48% kcal safflower oil plus 12% fish oil (n-3)] diets were fed to rats for 4 wk. After the dietary treatments, muscle lipid turnover and oxidation in the presence and absence of leptin was measured using pulse-chase procedures in incubated resting soleus muscle. In the absence of leptin, phospholipid, diacylglycerol, and triacylglycerol (TG) turnover were unaffected by the high-fat diets, but exogenous palmitate oxidation was significantly increased in the HF-Saff group. In Cont rats, leptin increased exogenous palmitate oxidation (21.4 +/- 5.7 vs. 11.9 +/- 1.61 nmol/g, P = 0.019) and TG breakdown (39.8 +/- 5.6 vs. 27.0 +/- 5.2 nmol/g, P = 0.043) and decreased TG esterification (132.5 +/- 14.6 vs. 177.7 +/- 29.6 nmol/g, P = 0.043). However, in both high-fat groups, the stimulatory effect of leptin on muscle lipid oxidation and hydrolysis was eliminated. Partial substitution of fish oil resulted only in the restoration of leptin's inhibition of TG esterification. Thus we hypothesize that, during the development of obesity, skeletal muscle becomes resistant to the effects of leptin, resulting in the accumulation of intramuscular TG. This may be an important initiating step in the development of insulin resistance common in obesity.

Endurance training increases FFA oxidation and reduces triacylglycerol utilization in contracting rat soleus.

We examined the effects of 8 wk of intense endurance training on free fatty acid (FFA) transporters and metabolism in resting and contracting soleus muscle using pulse-chase procedures. Endurance training increased maximal citrate synthase activity in red muscles (+54 to +91%; P </= 0.05) but failed to increase cytosolic fatty acid binding protein content, mRNA for fatty acyl-CoA synthase, and the putative FFA transporters or transport of palmitic acid into giant sarcolemmal vesicles. At rest, only triacylglycerol (TG) synthesis was significantly increased by training (+100.9 +/- 8.7 vs. +66.6 +/- 6.7 nmol/g wet wt; P </= 0.05). Muscle contraction increased TG synthesis (+46%; P </= 0.05) and palmitate oxidation (+115%; P </= 0.05) in untrained rats. Endurance training further enhanced synthesis of monoacylglycerol (MG), diacylglycerol (DG) and TG during contraction (+36, +69 and +71%, respectively; P </= 0.05), as well as exogenous palmitate oxidation (+41%; P </= 0.05) relative to untrained rats. Compared with those in untrained rats, TG breakdown and oxidation during contraction were reduced after training by 49 and 30%, respectively (P </= 0.05). In conclusion, endurance training 1) increases FFA oxidation and incorporation into endogenous lipid pools during contraction and 2) reduces the rate of intramuscular TG utilization during contraction when exogenous FFA availability is adequate. The enhanced FFA uptake subsequent to training appears to be independent of altered maximal transport rates of FFA into the muscle cell.

Structural and Biochemical Changes in Loblolly Pine (Pinus taedaL.) Seeds during Germination and Early Seedling Growth. II. Storage Triacylglycerols and Carbohydrates

Triacylglycerols (TAGs) comprised 59% of the total storage reserve in mature loblolly pine (Pinus taeda L.) seeds; 80% of these TAGs were stored in the megagametophyte. The TAG breakdown in the seedling was initiated before radicle emergence (during germination), while in the megagametophyte breakdown occurred after radicle emergence (during early seedling growth). In both seed tissues, the majority of TAG breakdown took place during early seedling growth. Within the seedling, the most rapid rate of TAG breakdown occurred in the radicle and hypocotyl. Unlike TAGs, there was very little carbohydrate stored in loblolly pine seeds at maturity. Levels of 80% ethanol‐soluble carbohydrate in the megagametophyte and seedling decreased during germination and then increased during early seedling growth. This increase coincided with the period of rapid TAG depletion. Accompanying the increase in 80% ethanol‐soluble carbohydrate level was a corresponding increase in 80% ethanol‐insoluble carbohydrates, such as starch, in the seedling during early seedling growth. Accumulation of starch occurred in both the cotyledons and hypocotyl. Starch accumulation in the megagametophyte was more transient, occurring around germination. The megagametophyte was important for the growth and nutrition of the seedling. In the presence of the megagametophyte, the seedling accumulated sucrose during early seedling growth. However, in the absence of the megagametophyte, loblolly pine seedlings failed to accumulate carbohydrates to any great extent.

Expression of hormone-sensitive lipase and its regulation by adrenaline in skeletal muscle

The enzymic regulation of triacylglycerol breakdown in skeletal muscle is poorly understood. Western blotting of muscle fibres isolated by collagenase treatment or after freeze-drying demonstrated the presence of immunoreactive hormone-sensitive lipase (HSL), with the concentrations in soleus and diaphragm being more than four times the concentrations in extensor digitorum longus and epitrochlearis muscles. Neutral lipase activity determined under conditions optimal for HSL varied directly with immunoreactivity. Expressed relative to triacylglycerol content, neutral lipase activity in soleus muscle was about 10 times that in epididymal adipose tissue. In incubated soleus muscle, both neutral lipase activity against triacylglycerol (but not against a diacylglycerol analogue) and glycogen phosphorylase activity increased in response to adrenaline (epinephrine). The lipase activation was completely inhibited by anti-HSL antibody and by propranolol. The effect of adrenaline could be mimicked by incubation of crude supernatant from control muscle with the catalytic subunit of cAMP-dependent protein kinase, while no effect of the kinase subunit was seen with supernatant from adrenaline-treated muscle. The results indicate that HSL is present in skeletal muscle and is stimulated by adrenaline via beta-adrenergic activation of cAMP-dependent protein kinase. The concentration of HSL is higher in oxidative than in glycolytic muscle, and the enzyme is activated in parallel with glycogen phosphorylase.

Effects of epinephrine on lipid metabolism in resting skeletal muscle.

The effects of physiological (0, 0.1, 2.5, and 10 nM) and pharmacological (200 nM) epinephrine concentrations on resting skeletal muscle lipid metabolism were investigated with the use of incubated rat epitrochlearis (EPT), flexor digitorum brevis (FDB), and soleus (SOL) muscles. Muscles were chosen to reflect a range of oxidative capacities: SOL > EPT > FDB. The muscles were pulsed with [1-14C]palmitate and chased with [9,10-3H]palmitate. Incorporation and loss of the labeled palmitate from the triacylglycerol pool (as well as mono- and diacylglycerol, phospholipid, and fatty acid pools) permitted the simultaneous estimation of lipid hydrolysis and synthesis. Endogenous and exogenous fat oxidation was quantified by 14CO2 and 3H2O production, respectively. Triacylglycerol breakdown was elevated above control at all epinephrine concentrations in the oxidative SOL muscle, at 2.5 and 200 nM (at 10 nM, P = 0.066) in the FDB, and only at 200 nM epinephrine in the EPT. Epinephrine stimulated glycogen breakdown in the EPT at all concentrations but only at 10 and 200 nM in the FDB and had no effect in the SOL. We further characterized muscle lipid hydrolysis potential and measured total hormone-sensitive lipase content by Western blotting (SOL > FDB > EPT). This study demonstrated that physiological levels of epinephrine cause measurable increases in triacylglycerol hydrolysis at rest in oxidative but not in glycolytic muscle, with no change in the rate of lipid synthesis or oxidation. Furthermore, epinephrine caused differential stimulation of carbohydrate and fat metabolism in glycolytic vs. oxidative muscle. Epinephrine preferentially stimulated glycogen breakdown over triacylglycerol hydrolysis in the glycolytic EPT muscle. Conversely, in the oxidative SOL muscle, epinephrine caused an increase in endogenous lipid hydrolysis over glycogen breakdown.

Cyclic AMP and fatty acids increase carnitine palmitoyltransferase I gene transcription in cultured fetal rat hepatocytes.

In the rat, the gene for liver mitochondrial carnitine palmitoyltransferase I (CPT I), though dormant prior to birth, is rapidly activated postnatally. We sought to elucidate which hormonal and/or nutritional factors might be responsible for this induction. In cultured hepatocytes from 20-day-old rat fetus, the concentration of CPT I mRNA, which initially was very low, increased dramatically in a dose-dependent manner after exposure of the cells to dibutyryl cAMP (Bt2cAMP). Similar results were obtained when long-chain fatty acids (LCFA), but not medium-chain fatty acids, were added to the culture medium. The effects of Bt2cAMP and LCFA were antagonized by insulin, also dose dependently. In contrast, CPT II gene expression, which was already high in fetal hepatocytes, was unaffected by any of the above manipulations. Bt2cAMP stimulated CPT I gene expression even when endogenous triacylglycerol breakdown was suppressed by lysosomotropic agents suggesting that the actions of cAMP and LCFA were distinct. Moreover, half-maximal concentrations of Bt2cAMP and linoleate produced an additive effect CPT I mRNA accumulation. While linoleate and Bt2cAMP stimulated CPT I gene transcription by twofold and fourfold, respectively, the fatty acid also increased the half-life of CPT I mRNA (50%). When hepatocytes were cultured in the presence of 2-bromopalmitate, (which is readily converted by cells into its non-metabolizable CoA ester) CPT I mRNA accumulation was higher than that observed with oleate or linoleate. Similarly, the CPT I inhibitor, tetradecylglycidate, which at a concentration of 20 microM did not itself influence the CPT I mRNA level, enhanced the stimulatory effect of linoleate. The implication is that induction of the CPT I message by LCFA does not require mitochondrial metabolism of these substrates; however, formation of their CoA esters is a necessary step. Unlike linoleate, the peroxisome proliferator, clofibrate, increased both CPT I and CPT II mRNA levels and neither effect was offset by insulin. It thus appears that the mechanism of action of LCFA differs from that utilized by clofibrate, which presumably works through the peroxisome proliferator activated receptor. We conclude that the rapid increase in hepatic CPT I mRNA level that accompanies the fetal to neonatal transition in the rat is triggered by the reciprocal change in circulating insulin and LCFA concentrations, coupled with elevation of the liver content of cAMP.

Effects of oxfenicine on the atria from fed and fasted rats.

The aim of the investigation was to assess whether endogenous triacylglycerol contributes to the maintenance of the atrial functions. To attain this information, the atria from fed and fasted rats were treated with oxfenicine which is a cardioselective inhibitor of carnitine palmitoyltransferase I. In the presence of glucose, oxfenicine suppressed lipolysis without affecting the pacemaker and contractile activities. When exposed to 2-deoxyglucose in a substrate-free medium, the atria displayed a progressive fall of the contractile strength and pacemaker rate. The dysfunctions appeared faster in the atria from fed rats coinciding with a smaller triacylglycerol mobilization. Under this condition, oxfenicine abolished the triacylglycerol breakdown, increased the fall in the peak tension, elicited a rise in the resting tension and accelerated the decline of the pacemaker rate, leading in a significant number of atria to a complete cessation of the spontaneous contractions. These effects proceeded faster in the fed rats atria. Present data suggest that glucose oxidation is sufficient to meet the atrial energy demand when the fatty acid catabolism is impeded. The noxious effects of oxfenicine, attained after the glucose metabolism was eliminated, lend direct evidence to the notion that endogenous triacylglycerol supports, at least partly, the atrial functions.

Osmotic potential and abscisic acid regulate triacylglycerol synthesis in developing wheat embryos

This work was carried out to determine what factors in the developing wheat (Triticum aestivum L.) grain are involved in regulating the metabolism of the triacylglycerol (TAG) storage reserves. When embryos are isolated from the grain and incubated in media for 4 d the TAG content is affected in three ways. In the basal medium (dilute buffer) the content falls; in 30 mM sucrose the content remains unchanged; in sucrose supplemented with an osmoticum (400 mM mannitol) or abscisic acid (1 μM ABA) the TAG content increases. Effective osmotic potentials and ABA concentrations fit well with their respective values in planta. The fatty-acid composition of TAG accumulated in vitro is close to that in planta but in the absence of ABA or osmoticum there is a fall in the C18∶C16 ratio. Experiments with [14C] acetate show that the in-planta rate of incorporation into TAG can only occur in isolated embryos treated osmotically or with ABA, while there seems to be no effect of these two factors on TAG breakdown. An osmotic ‘shock’ (dilute buffer) for only 2 h causes a rapid fall in TAG synthesis which continues for ca. 24 h after which it recovers. Abscisic acid protects against osmotic ‘shock’. It is concluded that TAG synthesis in developing wheat embryos is regulated by the osmotic potential and-or ABA, and that the embryos are very sensitive to short-term perturbations of these two factors.

Deacylation and uptake of the fatty acyl chain from enantiomeric dialkylacylglycerol by intact rat fat cells

Isolated rat fat cells hydrolyze the ester bond in position 3 of enantiomeric dialkyl[ 3H]acylglycerol added as triester-analogue substrate. This lipolytic system constitutes an integrated model which allows, in the absence of di- and monoesters, specific determinations of both the rate-limiting step in triacylglycerol breakdown and the uptake of acyl residues in cell lipids. Splitting of the ester bond was catalyzed by a cell-bound acylhydrolase which exhibited the characteristic properties of lipoprotein lipase. About 70% of the total amount of 3H-labelled fatty acid released during hydrolysis were concomitantly taken up by the cells and reesterified, mainly as triacylglycerol, in cell lipids. The acyl residues appeared to penetrate the cells directly after lipoprotein lipase hydrolysis, without entering the extracellular compartment. This, along with previous finding of a high positive correlation between rates of lipolytic activity and fatty acid uptake (Vérine A., Salers P. and Boyer J. (1982) Am. J. Physiol. 243, E175-E181), suggests that the uptake process is directly related to enzyme action. These results are consistent with the concept that lipoprotein lipase, besides being an acylhydrolase, could also function as acyltransferase.

Short-term regulation of hepatic triacylglycerol metabolism by insulin and glucagon.

This paper presents a review of our current knowledge of short-term regulation of triacylglycerol metabolism by insulin and glucagon in isolated rat hepatocytes. Insulin is devoted to rapidly improve the lipogenic climate of the hepatocyte. The hormone stimulates glycolysis, the conversion of pyruvate into acetyl-CoA, the synthesis of fatty acids and their esterification. The opposite pathways, hydrolysis of triacylglycerols and fatty acid oxidation, are depressed by insulin. Glucagon does all but deteriorate the lipogenic climate within the liver cell as the hormone inhibits triacylglycerol synthesis and stimulates the breakdown of triacylglycerols.

Cyclic AMP metabolism and lipolysis in cultured human adipocyte precursors

Triacylglycerol breakdown (lipolysis) results from a series of reactions culminated by activation of "hormone-stimulated" triacylglycerol lipase, an enzyme unique to adipose tissue. We have studied various components of the lipolytic process in human omental adipocyte precursors differentiating in culture. The levels of cyclic AMP, the "second messenger" of lipolytic hormones, were about sixfold higher in fat cell precursors than those in abdominal skin fibroblasts. L-Isoproterenol resulted in significant elevation of cyclic AMP levels in both cell types. Preincubation of intact adipocyte precursors with insulin resulted in significant enhancement of "low Km" cyclic AMP phosphodiesterase activity; in contrast, this hormone had no effect on fibroblast phosphodiesterase activity, a distinctive biochemical difference despite the morphological similarities between the two cell types during the early stages of adipocyte precursor maturation. Incubation of adipocyte precursors with isoproterenol resulted in the release of fatty acids into the medium, findings indicative of "hormone-stimulated" lipase activity and, hence, the operation of the entire "lipolytic cascade"; isoproterenol-stimulated lipolysis was inhibited by insulin. Release of fatty acids from fibroblasts was not observed. Thus, "hormone-stimulated" lipolysis and insulin stimulation of cyclic AMP phosphodiesterase activity are expressed during early stages of human adipocyte precursor differentiation.

Skeletal muscle triacylglycerol in the rat: methods for sampling and measurement, and studies of biological variability.

Previously reported concentrations of triacylglycerol in skeletal muscle have shown high coefficients of variation, and there have been large differences between mean concentrations reported in a given muscle. Conditions for sampling and measurement were therefore investigated. Samples were best taken under anesthesia as breakdown of triacylglycerol was rapid after decapitation. Silicic acid was preferable to zeolite for removal of phospholipids although either agent could interfere with the estimation. Even with apparently reliable methods, a high variability was found in any one muscle and there were large differences between muscles. It is unlikely that the variability was due to contamination with adipose tissue. Concentrations of glycogen and phospholipid were much less variable. Although the store of triacylglycerol in skeletal muscle in caloric terms was found to be 2-18 times greater than that of glycogen, the variability found is likely to hamper studies of its metabolic role.

Lipases of rat brain microsomes.

There exists a considerable degree of information in regard to the activities of glycerol ester hydrolases from numerous tissue sources. However, little attention has been given to the study of glyceride lipases in brain. Brain, in comparison to other organs, contains smaller quantities of neutral glycerides; amounts of triacylglycerol from whole rat brain have been established at 0.15–0.17 umol/g tissue (1). Numerous observations demonstrate that acylglycerols are metabolically active in brain tissue (2–5). In addition, several findings suggest the possibility for the presence of lipase activity in brain. These include the production of free fatty acids in incubated slices of mouse brain (6), and the rapid appearance and loss of radioactivity inthe triacylglycerols of mouse brain following intracerebral injection of (1-14C)oleic acid (7). Other observations report a reduction in the concentration of membrane diacylglycerol levels in crude homogenates of rat brain (8) and an increase in the amount of free fatty acids at the onset of brain ischemia (9). Ischemia was in turn accompanied by triacylglycerol breakdown, and the involvement of a separate diacylglycerol lipase was postulated (10).

Influence of α- dl-tocopherol on fungal lipid composition

The addition of α- dl-tocopherol to growing cultures of the fungus Glomerella cingulata increased triacylglycerol content 50% or more but decreased the synthesis of triacylglycerols approximately 16-fold; moreover, α-tocopherol treated cultures had 10% to 85% less malonaldehyde than control cultures. These results suggest that α-tocopherol caused increased triacylglycerol content by decreasing triacylglycerol breakdown. In contrast, phosphoglyceride content and synthesis remained virtually unchanged by the addition of α-tocopherol. In both classes of glycerolipid, treatment with α-tocopherol led to increased ratios of unsaturated to saturated fatty acids; but, α-tocopherol treatment led to an increase in the number of double bonds per mole of fatty acid in the triacylglycerols and to a decrease in the number of double bonds per mole of fatty acid in the phosphoglycerides. We infer that, through deacylation-reacylation reactions, the triacylglycerols of treated cultures supplied the phosphoglycerides with relatively saturated fatty acids and that the triacylglycerols also were used for storage of relatively unsaturated fatty acids derived from the phosphoglycerides. Our results suggest that α-tocopherol can be used to produce predictable shifts in the fatty acid composition of fungal phosphoglycerides. Such shifts can be useful tools in attempting to understand how changes in the fatty acid composition of phospholipids influence the physical properties and functions of cellular membranes.

Triacylglycerol turnover in Tetrahymena pyriformis relation to phospholipid synthesis

The metabolic function of triaclyglycerol in Tetrahymena pyriformis was investigated by prelabeling endogenous lipid with a 14C-labeled short chain fatty acid, and then following the disappearance of radioactivity from triacylglycerol and its appearance in other products. In 90 min, up to 85% of the label in triacylglycerol turns over, and although some radioactivity appears in CO 2 and glycogen, most of the label appears in phospholipid. Starvation of the cells, as well as resuspension in enriched medium or provision of acetate all block triacylglycerol breakdown, while supplementation of the medium with pyruvate does not. Prelabeling lipid with [ 3H]glycerol shows that some of the transfer of material from triacylglycerol to phospholipid involves transfer of the glycerol backbone, although transfer of triacylglycerol fatty acids directly to phospholipid probably also occurs. In addition, the catabolism of triacylglycerol occurs by a “last-in-first-out” mechanism, indicating some form of compartmentation of triacylglycerol in this cell. The results demonstrate an important metabolic interrelationship between triacylglycerol catabolism and phospholipid synthesis and raise the question, in this cell at least, of the validity of considering triacylglycerol only as a fuel storage form.