Regulation of biosynthesis of secondary metabolites

The rate of fatty acid synthesis withStreptomyces aureofaciens was measured by means of incorporation of 1-¹⁴C-acetic acid. The maximum specific rate of incorporation was achieved in the 12th hour. The incorporation rate of the labelled acetate into chlortetracycline was roughly twice that into fatty acids during the production phase. The rate of fatty acid synthesis in the high producing strain was five times that of the low producing one, although the dry weight fatty acid content was almost the same in both the strains.     

Effect of Ethrel and Ceratocystis fimbriata on the Synthesis of Fatty Acids and 6-Methoxy Mellein in Carrot Root

The rate of incorporation of ¹⁴C from acetate-1-¹⁴C into fatty acids by carrot root discs, 18 hours after inoculation with Ceratocystis fimbriata, was 9-fold greater than that in freshly cut discs. The rate in discs treated with water or Ethrel was 3-fold greater. The rate of incorporation of ¹⁴C from glucose-U-¹³C into fatty acids was 3-fold greater 18 hours after any of the above treatments. The rate of ¹⁴C incorporation from malonate-2-¹⁴C into fatty acids 24 hours after inoculation with C. fimbriata or treatment with water was 25 and 60%, respectively, of that in freshly cut discs. Linoleic acid was the principal fatty acid in carrot root, but incorporation of ¹⁴C from acetate-1-¹⁴C into the acid was low until 18 hours after inoculation with C. fimbriata or treatment with Ethrel. Turnover rates of the fatty acids appeared low and were similar for all treatments.     

Metabolism of tween-Fatty Acid esters by cultured soybean cells : kinetics of incorporation into lipids, subsequent turnover, and associated changes in endogenous Fatty Acid synthesis

Uptake of Tween-fatty acid esters and incorporation of the fatty acids into lipids by soybean (Glycine max [L.] Merr.) suspension cultures was investigated, together with subsequent turnover of the incorporated fatty acids and associated changes in endogenous fatty acid synthesis. Tween uptake was saturable, and fatty acids were rapidly transferred from Tweens to all acylated lipids. Patterns of incorporation into glycerolipids were similar in cells treated with Tweens carrying [1-¹⁴C]-fatty acids and in cells treated with [1-¹⁴C]acetate, indicating that exogenous fatty acids were used for glycerolipid synthesis essentially as if they had been made by the cell. In Tween-treated cells neutral lipids (which include Tweens) initially accounted for the majority of lipid radioactivity. Radioactivity was then rapidly transferred to glycerolipids. A transient pool of free fatty acids accounting for up to 10% of lipid radioactivity was observed. This was consistent with the hypothesis that fatty acids are transferred from Tweens to lipids by deacylation of the Tweens, creating a pool of free fatty acids which are then used for lipid synthesis. Sterols were only slightly labeled in cells treated with Tweens, but accounted for nearly 50% of lipid radioactivity in cells treated with acetate. This suggested very little degradation and reutilization of the radioactive fatty acids in cells treated with Tweens. In cells treated with either [1-¹⁴C]acetate or Tween-[1-¹⁴C]-18:1, 70% of the initial fatty acid radioactivity remained in fatty acids after a 100 hour chase. By contrast, fatty acids not normally present disappeared more rapidly, suggesting differential treatment of such fatty acids compared with those normally present. Cells which had incorporated large amounts of exogenous fatty acids altered fatty acid synthesis in three distinct ways: (a) amounts of [1-¹⁴C]acetate incorporated into fatty acids were reduced; (b) cells incorporating exogenous unsaturated fatty acids increased the proportion of [1-¹⁴C]acetate partitioned into saturated fatty acids, while the converse was true of cells which had incorporated exogenous saturated fatty acids; (c) desaturation of 18:1 to 18:2 and 18:3 was reduced in cells which had incorporated unsaturated fatty acids. These results suggest that Tween-fatty acid esters will be useful for supplying fatty acids to cells for a variety of studies related to fatty acid or membrane metabolism.     

A COMPARATIVE STUDY OF THE METABOLISM OF DE NOVO SYNTHESIZED FATTY ACIDS FROM ACETATE AND GLUCOSE, AND EXOGENOUS FATTY ACIDS, IN SLICES OF RABBIT CEREBRAL CORTEX DURING DEVELOPMENT

Slices of rabbit cerebral cortex, from the foetal stage to the adult have been used to compare lipid synthesis from fatty acids synthesized de novo from [U-¹⁴C]glucose and [1-¹⁴C]acetate, with lipid synthesis from exogenous albumin-bound [1-¹⁴C]palmitate. Incorporation into cellular lipid has been determined in terms of DNA, protein, wet wt. of tissue and wet weight of whole brain. On a wet wt. basis, maximum incorporation of glucose carbon into lipid occurred in the foetal brain while lipid synthesis from acetate and palmitate was maximum at 4–14 days after birth. Glucose and acetate were incorporated into a diversity of lipids (with increasing amounts of phosphatidylcholine synthesized during maturation), while palmitate was incorporated into the free fatty acid and triglyceride fractions. A greater proportion of acetate was incorporated into fatty acids of chain-length longer than C16 compared with the incorporation of palmitate. However, on a molar basis de novo synthesized and exogenous palmitate were elongated, desaturated and incorporated into phospholipids at a similar rate, while exogenous palmitate was incorporated to a greater extent than de nova synthesized fatty acid into the triglyceride fraction. This difference in metabolism may be due to the different size of the non-esterified fatty acid pool in the two situations. At the period of their most active formation, the very long-chain fatty acids may be synthesized from a pool of the C18 series of fatty acids (saturated and monoenoic) not in equilibrium with the bulk of C₁₈ acids in cerebral lipids. This could be a pool of acyl groups derived from ethanolamine phospholipids.     

Preincubation stimulates fatty acid synthesis by liver slices and isolated hypatocytes from fasted and diabetic rats.

We have observed that preincubation of 48 hour-fasted or alloxan diabetic rat liver slices, with no exogenous energy supply, for 3 hours resulted in an increased rate of incorporation of [1-¹⁴C] acetate into fatty acids and cholesterol during the following 2 hours. This preincubation effect was enhanced by the presence of glucose (25mM) in or prevented by the addition of dibutyryl cyclic adenosine 3′,5′ monophosphate (10^?4M) to the preincubation medium. Preincubation of normal rat liver slices did not change their rate of incorporation of [1-¹⁴C] acetate into fatty acids or cholesterol. The rate of ¹⁴CO₂ synthesized by normal, fasted or diabetic liver slices was little affected by preincubation. The preincubation effect, i.e. enhanced fatty acid synthesis was also observed in suspensions of hepatocytes from fasted and diabetic rats, preincubated for 2 hours, followed by a 1 hour incubation with either [1-¹⁴C] acetate or [³H] H₂O as precursor. We conclude from these data that there is concurrent and coordinated short- and long-term regulation of fatty acid biosynthesis in fasted and diabetic rat livers. Further, we suggest that the release of inhibition by preincubation of these tissues provides a useful tool for studying the coordinated control     

The effects of specific lipogenic substrates and metabolic inhibitors on de Novo fatty acid synthesis in isolated hepatocytes from chow-fed female rats

The effects of glucose (10 mm), glycerol (3 mm), and lactate/pyruvate (10 mm) on the incorporation of ³H from ³H₂O into fatty acids were studied in isolated hepatocytes prepared from chow-fed female rats. Lactate/pyruvate markedly increased lipogenic rates, while glucose and glycerol did not significantly affect rates of lipogenesis. In cells incubated with lactate/pyruvate plus glycerol, the increase in ³H incorporation was greater than observed with lactate/pyruvate alone. In hepatocytes isolated from 24-h starved rats, lactate/pyruvate again increased de novo fatty acid synthesis to a greater extent than either glucose or glycerol. Glycerol significantly increased lipogenesis compared to the endogenous rates and when incubated with lactate/pyruvate produced an increase above lactate/pyruvate alone. (?)-Hydroxycitrate, a potent inhibitor of ATP-citrate lyase (EC 4.1.3.8), and agaric acid, an inhibitor of tricarboxylate anion translocation, were studied in hepatocytes to determine their effects on lipogenesis by measuring ³H₂O, [1-¹⁴C]acetate, and [2-¹⁴C]lactate incorporation into fatty acids. ³H incorporation into fatty acids was markedly inhibited by both inhibitors with agaric acid (60 μm) producing the greater inhibition. (?)-Hydroxycitrate (2 mm) increased acetate incorporation into fatty acids from [1-¹⁴C]acetate and agaric acid produced a strong inhibitory effect. Combined effects of (?)-hydroxycitrate and agaric acid on lipogenesis from [1-¹⁴C]acetate showed an inhibitory response to a lesser extent than with agaric acid alone. With substrate concentrations of acetate present, there was no significant increase in rates of lipogenesis from [1-¹⁴C]acetate and the increase previously observed with (?)-hydroxycitrate alone was minimized. Agaric acid significantly inhibited fatty acid synthesis from acetate in the presence of exogenous substrate, but the effect was decreased in comparison to rates with only endogenous substrate present. With [2-¹⁴C]lactate as the lipogenic precursor, agaric acid and (?)-hydroxycitrate strongly inhibited fatty acid synthesis. However, agaric acid despite its lower concentration (60 μm vs 2 mm) was twice as effective as (?)-hydroxycitrate. A similar pattern was observed when substrate concentrations of lactate/pyruvate (10 mm) were added to the incubations. When (?)-hydroxycitrate and agaric acid were simultaneously incubated in the presence of endogenous substrate, there was an additive effect of the inhibitors on decreasing fatty acid synthesis. Results are discussed in relation to the origin of substrate for hepatic lipogenesis and whether specific metabolites increase lipogenic rates.     

Substrate utilization for energy production and lipid synthesis in oligodendrocyte-enriched cultures prepared from rat brain

The metabolism of oligodendrocytes has been studied using cultures of oligodendrocyte-enriched glial cells isolated from cerebra of 5–8-day old rats. Cultures containing 60–80% oligodendrocytes were incubated for 16h with [3-¹⁴C]acetoacetate, d-[3-¹⁴C]3-hydroxybutyrate, [U-¹⁴C]glucose, l-[U-¹⁴C]glutamine and [1-¹⁴C]pyruvate or [2-¹⁴C]pyruvate in the presence or absence of other oxidizable substrates. Labelled CO₂ was collected as an index of oxidative metabolism and the incorporation of label into total lipids, fatty acids and cholesterol was used as an index of the de novo synthesis of lipids. Glucose, acetoacetate, D-3-hydroxybutyrate, pyruvate and l-lactate were measured to determine substrate utilization and product formation under various conditions. Our results indicate that glucose is rapidly converted to lactate and is a relatively poor substrate for oxidative metabolism and lipid synthesis. Ketone bodies were used as an energy source and as precursors for the synthesis of fatty acids and cholesterol. Preferential incorporation of acetoacetate into cholesterol was not observed. Exogenous pyruvate was incorporated into both the glycerol skeleton of complex lipids and into cholesterol and fatty acids. l-Glutamine appeared to be an important substrate for the energy metabolism of these cells.     

Synthesis of medium-chain fatty acids and their incorporation into triacylglycerols by cell-free fractions fromCuphea embryos

During their rapid maturation period, seeds of Cuphea wrightii A. Gray mainly accumulate medium-chain fatty acids (C₈ to C₁₄) in their storage lipids. The rate of lipid deposition (40–50 mg·d^–1·(g fresh weight)^–1) is fourfold higher than in seeds of Cuphea racemosa (L. f.) Spreng, which accumulate long-chain fatty acids (C₁₆ to C₁₈). Measurements of the key enzymes of fatty-acid synthesis in cell-free extracts of seeds of different maturities from Cuphea wrightii show that malonyl-CoA synthesis may be a triggering factor for the observed high capacity for fatty-acid synthesis. Experiments on the incorporation of [1-¹⁴C]acetate into fatty acids by purified plastid preparations from embryos of Cuphea wrightii have demonstrated that the biosynthesis of medium-chain fatty acids (C₈ to C₁₄) is localized in the plastid. Thus, in the presence of cofactors for lipid synthesis (ATP, NADPH, NADH, acyl carrier protein, and sn-glycerol-3-phosphate), purified plastid fractions predominantly synthesized free fatty acids, 30% of which were of medium chain length. Transesterification of the freshly synthesized fatty acids to coenzyme A and recombination with the microsomal fraction of the embryo homogenate induced triacylglycerol synthesis. It also stimulated fatty-acid synthesis by a factor 2–3 and increased the relative amount of medium-chain fatty acids bound to triacylglycerols, which corresponded to about 60–80% in this lipid fraction.Abbreviations ACP acyl carrier protein - FW fresh weight This work was supported by the Bundesminister für Forschung und Technologie. The authors thank S. Borchert for her suggestions for plastid preparation.     

Nature of DNA and RNA Degradation in Escherichia coli Induced by Colicin E2

Lipogenesis in the fatty liver of rat, which was induced by feeding an amino acid unbalanced diet containing 8% casein supplemented with 0.3% dl-methionine, has been studied by measuring the incorporation of glycerol-1-¹⁴C, palmitate-1-¹⁴C, citrate-1,5-¹⁴C, pyruvate-1-¹⁴C and pyruvate-2-¹⁴C into various lipid fractions and ¹⁴CO₂ during in vitro incubation of liver slices.

The total radioactivity of liver lipid per 100 g of the body and the incorporation of each substrate into triglyceride in the lipid were significantly higher in the imbalance group than the control group. Conversion of each substrate to ¹⁴CO₂ was not imparied in the imbalance group.

It is evident from these results that the induction of this type of fatty liver is due mainly to the triglyceride synthesis by both the fatty acid synthesis and the transesterification of fatty acid.

These results are considered to support the previous assumption in which acetate-1-¹⁴C was used as a precursor.     

Incorporation of [214C]malonyl CoA into fatty acids by a cell-free extract of Catharanthus roseus suspension culture cells

A cell-free extract containing the enzymes for de-novo synthesis, elongation and desaturation of fatty acids was prepared from cultured cells of Catharanthus roseus G. Don. ¹⁴C-Fatty acids synthesized by the extract from [2-¹⁴C]malonyl CoA substrate were palmitic (16:0), stearic (18:0) and oleic (18:1). Dialyzed extract was active and stable at room temperature and at 4° C, but was inactivated on boiling. There was an absolute requirement for NADPH for incorporation of [2-¹⁴C]malonyl CoA into total fatty acids. Escherichia coli acyl carrier protein stimulated total fatty-acid synthesis without affecting the relative ratio of individual fatty acids. Total fatty-acid synthesis at a rate of 45 nmol·mg^-1 protein·h^-1 occurred at a substrate level of 73 M malonyl CoA, cofactor levels of 500 M NADPH, 30 g·ml^-1 E. coli ACP, and 1.0 mg·ml^-1 extract protein. Total fatty acid synthesis was also sensitive to cerulenin and CoA levels. Variations in the relative abundance of individual ¹⁴C-fatty acids were regulated by concentrations of [¹⁴C]malonyl CoA. NADPH and ferredoxin, as well as by pH, temperature and length of incubation. Fatty-acid synthetase enzymes responsible for [¹⁴C]palmitic acid were rapidly saturated at a low substrate level (0.3 M malonyl CoA). Increasing the level of [2-¹⁴C]malonyl CoA permitted further synthesis of [¹⁴C]stearate and [¹⁴C]oleate. Desaturation of [¹⁴C]stearate to [¹⁴C]oleate was stimulated by increasing the levels of NADPH and ferredoxin. The desaturase and elongase enzymes were sensitive to acidic pH. The desaturase was also unstable at 41° C, although fatty acid synthetase and elongase were unaffected by this temperature.Abbreviation ACP Acyl carrier protein     

THE INCORPORATION OF [1-14C]ACETATE INTO UNESTERIFIED FATTY ACIDS IN RAT CEREBRAL CORTEX IN VIVO

—The incorporation of [1-¹⁴C]acetate into unesterified fatty acids and into the fatty acids of neutral glycerides and of phospholipids has been measured in rat cerebral cortex in vivo. The most rapid incorporation is seen in the unesterified fatty acids which have a turnover time of 5-6 min. It is suggested that unesterified fatty acids are precursors to neutral glycerides and phospholipids rather than being derived from them by lipase activity.     

Comparison of acetate- and pyruvate-dependent fatty-acid synthesis by spinach chloroplasts

In recent studies using intact chloroplasts of spinach (Spinacia oleracea L.) to investigate the accumulation of acetyl-CoA produced by the activity of either acetyl-CoA synthetase (EC 6.2.1.1) or the pyruvate-dehydrogenase complex, this product was not detectable. These results in combination with new information on the physiological levels of acetate and pyruvate in spinach chloroplasts (H.-J. Treede et al. 1986, Z. Naturforsch. 41 C, 733–740) prompted a reinvestigation of the incorporation of [1-¹⁴C] acetate and [2-¹⁴C] pyruvate into fatty acids at physiological concentrations.The K _m for the incorporation into fatty acids was about 0.1 mM for both metabolites and thus agreed with the values obtained by H.-J. Treede et al. (1986) for acetyl-CoA synthetase and the pyruvate dehydrogenase complex. However, acetate was incorporated with a threefold higher V _max. Saturation for pyruvate incorporation into the fattyacid fraction was achieved only at physiological pyruvate concentrations (<1.0 mM). The diffusion kinetics observed at higher concentrations may be the result of contamination with derivates of the labeled substrate. Competition as well as double-labeling experiments with [³H]acetate and [2-¹⁴C]pyruvate support the notion that, at least in spinach, chloroplastic acetate is the preferred substrate for fatty-acid synthesis when both substrates are supplied concurrently (P.G. Roughan et al., 1979 b, Biochem. J. 184, 565–569).Experiments with spinach leaf discs confirmed the predominance of fatty-acid incorporation from acetate. Radioactivity from [1-¹⁴C]acetate appeared to accumulate in glycerolipids while that from [2-¹⁴C]pyruvate was apparently shifted in favor of the products of prenyl metabolism.Abbreviations Chl chlorophyll - TLC thin-layer chromatography     

Specific inhibition of candicidin biosynthesis by the lipogenic inhibitor cerulenin

Cerulenin, an inhibitor of fatty acid synthesis, inhibits specifically the biosynthesis of the polyene macrolide candicidin by resting cells of Streptomyces griseus. 50% inhibition was achieved with a cerulenin concentration of 1.5 μg/ml. Cells in which candicidin synthesis was inhibited for 10 h remained capable of candicidin synthesis after removal of the inhibitor. Cerulenin inhibits specifically the incorporation of [¹⁴C] propionate into candicidin but does not affect total protein or RNA synthesis. The uptake of [¹⁴C] propionate was not inhibited under conditions which totally prevented the incorporation of propionate into candicidin. Incorporation of p-amino[¹⁴C] benzoic acid NH₂[¹⁴C]BzO⁻ into the aromatic moiety of candicidin was also inhibited by cerulenin. The inhibitory action of cerulenin was not reversed by exogenous fatty acids. Since cerulenin is known to block the condensation of malonyl-CoA subunits in the formation of fatty acids, it is concluded that the polyene macrolide candicidin is synthesized via the polyketide pathway by condensation steps similar to those occurring in fatty acid biosynthesis.     

Fatty acid synthesis by slices from developing leaves

Fatty acid synthesis was studied in successive leaf sections from the base to the tip of developing barley (Hordeum vulgare L.), maize (Zea mays L.), rye grass (Lolium perenne L.) and wheat (Triticum aestivium L.) leaves. The basal regions of the leaves had the lowest rates of fatty acid synthesis and accumulated small amounts of very long chain fatty acids. Fatty acid synthesis was highest in the middle leaf sections in all four plants. Linolenic acid synthesis from [1-¹⁴C]acetate was highest in the distal leaf sections of rye grass. The labelling of the fatty acids of individual lipids of rye grass was examined and it was found that [¹⁴C]linolenic acid was highest in the galactolipids. Synthesis of this acid in the galactolipids was most active in leaf segment C. Only traces of [¹⁴C]linolenic acid were ever found in phosphatidylcholine and it is concluded that this phospholipid cannot serve as a substrate for linoleic acid desaturation in rye grass. The synthesis of fatty acids was sensitive to arsenite, fluoride and the herbicide EPTC. The latter was only inhibitory towards those leaf segments which made very long chain fatty acids. Formation of fatty acids from [1-¹⁴C]acetate was also studied in chloroplasts prepared from successive leaf sections of rye grass. Chloroplasts isolated from the middle leaf sections had the highest activity. Palmitic and oleic acids were the main fatty acid products in all chloroplast preparations. Linolenic acid synthesis was highest in chlorplasts isolated from the distal leaf sections of rye grass.     

Omega oxidation of fatty acids and the pathway of 3-hydroxybutyric acid formation

Pathways followed by the carbons of long chain fatty acids in their conversion to 3-hydroxybutyric acid were traced and the contribution of ω-oxidation to fatty acid oxidation was determined in the cellular environment where ketone body formation occurs. 1-¹⁴C-, 2-¹⁴C-, and ω-¹⁴C-labeled fatty acids were injected into alloxan-induced diabetic rats in ketosis. 3-Hydroxybutyric acid was isolated from their urines and degraded. About 1.2 to 1.4 times as much ¹⁴C was found in carbon 1 as carbon 3 of 3-hydroxybutyric acid when the 1-¹⁴C-labeled fatty acids were injected and in carbon 2 as carbon 4 when the 2-¹⁴C-labeled fatty acids were injected. There was about 4 times as much incorporation into carbon 4 as carbon 2 of 3-hydroxybutyric acid formed from the ω-¹⁴C-labeled fatty acids. This means that 50% or more of the fatty acids were oxidized, so that the terminal two carbons of the fatty acids were converted to acetoacetyl-CoA without acetyl-CoA as an intermediate. Incorporation of ¹⁴C into carbons 1 and 2 of the hydroxybutyric acid reflects the distribution of ¹⁴C in acetyl-CoA. Incorporation into carbon 1 was very small when the ω-¹⁴C-labeled fatty acids were substrate. This means that ω-oxidation of fatty acids makes, at most, a small contribution to the formation of the acetyl-CoA pool from which acetoacetate is derived.     

Ketone-body utilization and lipid synthesis by developing rat brain—a comparison between in vivo and in vitro experiments

The distribution of ketone bodies between oxidation and lipid synthesis was analysed in homogenates of developing rat brain. The capacity for lipid synthesis of homogenized or minced brain preparations was compared with rates of lipid synthesis in vivo, assessed by incorporation of ³H from ³H₂O into fatty acids and cholesterol. Brain homogenates of suckling rats (but not those of adults) incorporated label from [3-¹⁴C]ketone bodies into lipids, but this process was slow as compared to ¹⁴CO₂ production (< 5%) and much slower than the total rate of ketone-body utilization (< 0.5%). Study of ³H₂O incorporation demonstrated that the rates of lipogenesis and cholesterogenesis are at least one order of magnitude higher in vivo than in vitro. Maximal rates of ³H incorporation into fatty acids (3 μmol/g brain . h) and into cholesterol (0.6 μmol/g brain . h) were found during the third postnatal week. Adult rats still incorporated ³H into brain fatty acids at an appreciable rate (1 μmol/g brain . h), whereas cholesterogenesis was very low. It is concluded that in vitro measurements of lipid synthesis severely underestimate the rates that occur in developing rat brain in vivo. The high rate of ³H incorporation into lipids by developing and adult rat brain as compared to the amounts of these lipids present in the brain suggests an important contribution of endogenous lipid synthesis during brain development and an appreciable rate of fatty acid turnover during brain growth, but also in the adult brain.     

Relationship between fatty acid binding proteins,acetyl-CoA formation and fatty acid synthesis in developing human placenta

The relationship between fatty acid binding proteins, ATP citrate lyase activity and fatty acid synthesis in developing human placenta has been studied. Fatty acid binding proteins reverse the inhibitory efect of palmitoyl-CoA and oleate on ATP citrate lyase and fatty acid synthesis. In the absence of these inhibitors fatty acid binding proteins activate ATP citrate lyase and stimulate [ 1-¹⁴ C] acetate incorporation into placental fatty acids indicating binding of endogenous inhibitors by these proteins. Thus these proteins regulate the supply of acetyl-CoA as well as the synthesis of fatty acids from that substrates. As gestation proceeds and more lipids are required by the developing placenta fatty acid binding protein content, activity of ATP citrate lyase and rate of fatty acid synthesis increase indicating a cause and efect relationship between the demand of lipids and supply of precursor fatty acids during human placental development.     

Inhibition of hydrocarbon biosynthesis in the housefly by 2-Octadecynoate

The elongation of [9,10-³H]oleoyl-CoA with malonyl-CoA to form 20, 22, and 24 carbon monounsaturated fatty acids was demonstrated in housefly microsomes by radio-GLC. These elongation reactions, which have been postulated to be involved in hydrocarbon biosynthesis, have not been previously demonstrated in insects. 2-Octadecynoate (18:1 Δ²⁼) inhibited the in vivo incorporation of [1-¹⁴C]acetate into both fatty acids and hydrocarbons in a dose-dependent manner. At doses of 10 μg per female housefly of the alkynoic acid, the incorporation of [1-¹⁴C]acetate into hydrocarbon was inhibited 93%, the incorporation of [9,10-³H]oleate into hydrocarbon was inhibited 64%, and the incorporation of [1-¹⁴C]acetate into total internal lipid was inhibited 65%. Partially purified FAS was inhibited 50% and 95% at 15 μM and 40 μM, respectively, of the alkynoic acid. These results show that 2-octadecynoate inhibits hydrocarbon biosynthesis in the housefly by inhibiting FAS, and the in vivo data suggest that the elongation of 18:1 to longer chain fatty acids is also inhibited.     

Polyunsaturated Fatty Acid Biosynthesis in Cotyledons from Germinating and Developing Cucumis sativus L. Seedlings

Etiolated Cucumis sativus L. cotyledons preferentially catabolized exogenous [1-¹⁴C]oleic acid and [1-¹⁴C]linoleic acid with relatively little incorporation into complex lipids or desaturation of the ¹⁴C-labeled fatty acids. Following a 16-hour exposure to light, the greening cotyledons efficiently desaturated the exogenous ¹⁴C-labeled fatty acids. A small amount of oleate desaturation to linoleate was observed in etiolated tissue, but hardly any linoleate desaturation to α-linolenate was detected. Both oleate and linoleate desaturation showed diurnal variations with maxima at the end of light periods and minima at the end of dark periods. Illumination of etiolated tissue by flashing light, as opposed to continuous light, failed to stimulate either chlorophyll or α-linolenic acid biosynthesis, and both processes could be halted or reversed by 10 micrograms per milliliter cycloheximide. Production of polyunsaturated fatty acids from [1-¹⁴C]acetate, [1-¹⁴C]oleic acid, and [1-¹⁴C]linoleic acid, by greening cucumber cotyledons, was markedly affected by tissue integrity with finely chopped cotyledons having very little capacity for their synthesis and intact seedlings showing the highest rates.     

DEVELOPMENT OF LIPOGENESIS IN RAT BRAIN CORTEX: THE DIFFERENTIAL INCORPORATION OF GLUCOSE AND ACETATE INTO BRAIN LIPIDS IN VITRO

—The oxidation to CO₂ and the incorporation of [U-¹⁴C]glucose and [U-¹⁴C]acetate into lipids by cortex slices from rat brain during the postnatal period were investigated. The oxidation of [U-¹⁴C]glucose was low in 2-day-old rat brain, and increased by about two-fold during the 2nd and 3rd postnatal weeks. The oxidation of [U-¹⁴C]acetate was increased markedly in the second postnatal week, but decreased to rates observed in 2-day-old rat brain at the time of weaning. Both labeled substrates were readily incorporated into non-saponifiable lipids and fatty acids by brain slices from 2-day-old rat. Their rates of incorporation and the days on which maximum rates occurred were different, however, maximum incorporation of [U-¹⁴C]glucose and [U-¹⁴]acetate into lipid fractions being observed on about the 7th and 12th postanatal days, respectively. The metabolic compartmentation in the utilization of these substrates for lipogenesis is suggested. The activities of glucose-6-phosphate dehydrogenase, cytosolic NADP-malate dehydrogenase, cytosolic NADP-isocitrate dehydrogenase, ATP-citrate lyase and acetyl CoA carboxylase were measured in rat brain during the postnatal period. All enzymes followed somewhat different courses of development; the activity of acetyl CoA carboxylase was, however, the lowest among other key enzymes in the biosynthetic pathway, and its developmental pattern paralleled closely the fatty acid synthesis from [U-¹⁴C]glucose. It is suggested that acetyl CoA carboxylase is a rate-limiting step in the synthesis de novo of fatty acids in developing rat brain.