Control of fatty acid synthetase mRNA levels in rat liver by insulin, glucagon, and dibutyl cyclic AMP

The quantity of translatable fatty acid synthetase mRNA in liver of rats subjected to different hormonal states was determined with a rabbit reticulocyte lysate cell-free translation system. Both membrane-free polysomal and total cellular poly (A)-containing RNA were translated. The level of translatable fatty acid synthetase mRNA was 11-fold or more lower in livers of diabetic rats than in similar animals treated with insulin. In contrast, both glucagon and dibutyl cyclic AMP caused a 3-fold reduction over controls in the amount of translatable fatty acid synthetase mRNA in livers of animals refed a fat-free diet for 12 hr. These changes are consistent with the previously reported alterations in the relative rates of fatty acid synthetase synthesis measured in vivo. This suggests that the changes in the amount of fatty acid synthetase that occur in liver in response to the above hormonal changes are primarily due to changes in the amount of mRNA coding for this enzyme.     

Initiation of lipogenic enzyme activities in rat mammary glands.

The activities of acetyl-CoA carboxylase, ATP citrate-lyase and fatty acid synthetase remained low until parturition at 22 days of gestation and increased significantly within 1 day post partum. Administration of progesterone on days 20 and 21 and at parturition abolished the increases for at least 48 h after parturition. Removal of the pups of normal rats prevented the increases in activities of acetyl-CoA carboxylase and ATP citrate-lyase, but not of fatty acid synthetase, and administration of prolactin corticosterone or insulin did not stimulate activity. Tissue from suckled glands in which the ducts had been ligated at parturition showed no increase in the activities of acetyl-CoA carboxylase and ATP citrate-lyase within 24 h, whereas fatty acid synthetase activity was similar to that in the sham-operated contralateral glands. Foetoplacentectomy on day 18 increased the activity of fatty acid synthetase but not of acetyl-CoA carboxylase and ATP citrate-lyase; suckling of these dams by foster pups increased both acetyl-CoA carboxylase and ATP citrate-lyase.     

Regulation of the synthesis of ribulose-1,5-bisphosphate carboxylase and its subunits in the flagellate Chlorogonium elongatum. Different levels of translatable messenger RNAs for the large and the small subunits in autotrophic and heterotrophic cells as determined by immunological techniques

Poly(A)-rich and poly(A)-free RNAs were isolated from autotrophic and heterotrophic cells of the phytoflagellate Chlorogonium elongatum and translated in an mRNA-depleted reticulocyte lysate system. Immunoprecipitation methods were improved to detect large and small subunits of the chloroplast enzyme ribulose-1,5-bisphosphate carboxylase synthesized in vitro. Large-subunit polypeptides were shown to be the translation products of poly(A)-free RNA having the same molecular weight as large subunits made in vivo. Small-subunit polypeptides were synthesized when poly(A)-rich RNA was used as a template. They were made in vitro as a precursor, with an Mr about 6000 larger than mature small subunits. Cells growing heterotrophically in the dark with acetate are provided with lower levels of mRNA activities for the large and the small subunits is at least partially controlled by the amounts of translatable mRNAs.     

An improved purification and further characterization of the messenger ribonucleic acid for the fatty acid synthetase from rat liver

High purity fatty acid synthetase mRNA has been prepared from rat liver. The translational purity of the mRNA preparation was at least 27% as judged by the percentage of the radioactivity incorporated into acid-insoluble material that was precipitated by anti-fatty acid synthetase antibody. The specific activity of the mRNA was 220-times greater than that reported previously from this laboratory [1]. The large increase in the specific activity was achieved by the repeated use of high resolution linear-log sucrose density gradient centrifugation and the removal of 28 S rRNA by Sepharose 4B chromatography, as well as by the optimization of the K⁺ concentration (160 mM) in the reticulocyte lysate translation system. The mRNA preparation showed a single major band on agarose gel electrophoresis under denaturing conditions, and the translational activity of the fatty acid synthetase mRNA on the gel was found to coincide with this band. The molecular weight of the fatty acid synthetase mRNA is 2.5·10⁶ Da. The mRNA directed the synthesis of fatty acid synthetase with a molecular weight indistinguishable from that of the authentic enzyme subunit (M_r = 240 000). The copurification of the translation product and authentic enzyme revealed that the fatty acid synthetase polypeptides synthesized in the reticulocyte lysate system are assembled in vitro into dimers, the native form of the enzyme.     

Primary structure of chicken liver acetyl-CoA carboxylase deduced from cDNA sequence

The complete amino acid sequence of acetyl-CoA carboxylase from chicken liver has been deduced by cloning and sequence analysis of DNA complementary to its messenger RNA. The results were confirmed by Edman degradation of peptide fragments obtained by digestion of the enzyme polypeptide with Achromobacter proteinase I or staphylococcal serine proteinase. Chicken liver acetyl-CoA carboxylase is predicted to be composed of 2,324 amino acid residues, having a calculated molecular weight of 262,706. The biotin carboxyl carrier protein domain is located in the middle region of the enzyme polypeptide. The amino-terminal portion of the acetyl-CoA carboxylase has been found to exhibit a homologous primary structure to that of carbamyl phosphate synthetase. Localization of possible functional domains including biotin carboxylase subsite in the acetyl-CoA carboxylase polypeptide is discussed.     

The effect of progesterone on prolactin stimulation of fatty acid synthesis, glycerolipid synthesis and lipogenic-enzyme activities in mammary glands of pseudopregnant rabbits, after explant culture or intraductal injection.

The activities of lipogenic enzymes, such as acetyl-CoA carboxylase, fatty acid synthetase and glucose-6-phosphate dehydrogenase, and glycerolipid synthesis increased significantly in mammary explants of 11-day-pseudopregnant rabbits in response to prolactin, in the presence of near-physiological concentrations of insulin and corticosterone in culture. Increasing the concentration of progesterone in culture resulted in suppression of glycerolipid synthesis and activities of acetyl-CoA carboxylase and fatty acid synthetase, but not the pentose phosphate dehydrogenases. However, at near-physiological concentration of progesterone, only acetyl-CoA carboxylase activity was decreased. Injection of prolactin intraductally into 11-day-pseudopregnant rabbits stimulated glycerolipid synthesis, fatty acid synthesis and enzymes involved in fatty acid synthesis, after 3 days. Intraductal injection of progesterone separately or together with prolactin had no significant effect on basal or stimulated lipogenesis in mammary glands. Intramuscular injection of progesterone at 10 mg/day did not suppress fatty acid synthesis stimulated when prolactin was injected intraductally, but a significant inhibition was observed at a higher dose (80 mg/day).     

Adaptive synthesis of fatty acid synthetase and acetyl-CoA carboxylase by isolated rat liver cells.

Hepatocytes were isolated at specified times from livers of diabetic and insulin-treated diabetic rats during the course of a 48-h refeeding of a fat-free diet to previously fasted rats. The rates of synthesis of fatty acid synthetase and acetyl-CoA carboxylase in the isolated cells were determined as a function of time of refeeding by a 2-h incubation with l-[U-¹⁴C]leucine. Immunochemical methods were employed to determine the amount of radioactivity in the fatty acid synthetase and acetyl-CoA carboxylase proteins. The amount of radioactivity in the fatty acid synthetase synthesized by the isolated cells was also determined following enzyme purification of the enzyme to homogeneity. Enzyme activities of the fatty acid synthetase and acetyl-CoA carboxylase in the cells were measured by standard procedures. The results show that isolated liver cells obtained from insulintreated diabetic rats retain the capacity to synthesize fatty acid synthetase and acetyl-CoA carboxylase. The rate of synthesis of the fatty acid synthetase in the isolated cells was similar to the rate found in normal refed animals in in vivo experiments [Craig et al. (1972) Arch. Biochem. Biophys. 152, 619–630; Lakshmanan et al. (1972) Proc. Nat. Acad. Sci. USA69, 3516–3519]. In addition the relative rate of synthesis of fatty acid synthetase was stimulated greater than 20-fold in the diabetic animals treated with insulin. Immunochemical assays, when compared with enzyme activities, indicated the presence of an immunologically reactive, but enzymatically inactive, form or “apoenzyme” for both the fatty acid synthetase and acetyl-CoA carboxylase. The synthesis of these immunoreactive and enzymatically inactive species of protein, as well as the synthesis of the “holoenzyme” forms of both enzymes, requires insulin.     

Physiological regulation of acetyl-CoA carboxylase gene expression: effects of diet, diabetes, and lactation on acetyl-CoA carboxylase mRNA

We measured acetyl-CoA carboxylase mRNA levels in various tissues of the rat under different nutritional and hormonal states using a cDNA probe. We surveyed physiological conditions which are known to alter carboxylase activity, and thus fatty acid synthesis, to determine whether changes in the levels of carboxylase mRNA are involved. The present studies include the effects of fasting and refeeding, diabetes and insulin, and lactation on carboxylase mRNA levels. Northern blot analysis of liver RNA revealed that fasting followed by refeeding animals a fat-free (high carbohydrate) diet dramatically increased the amount of carboxylase mRNA compared to the fasted condition. These changes in the level of mRNA correspond to changes in the activity and amount of acetyl-CoA carboxylase. Acetyl-CoA carboxylase mRNA levels in epididymal fat tissue decreased upon fasting and increased to virtually normal levels after 72 h of refeeding, closely resembling the liver response. The amount of acetyl-CoA carboxylase mRNA decreased markedly in epididymal fat tissue of diabetic rats as compared to nondiabetic animals. However, 6 h after injection of insulin the mRNA level returned to that of the nondiabetic animals. Gestation and lactation also affected the levels of carboxylase mRNA in both liver and mammary gland. Maximum induction in both tissues occurred 5 days postpartum. These studies suggest that these diverse physiological conditions affect fatty acid synthesis in part by altering acetyl-CoA carboxylase gene expression.     

Translation and characterization of the fatty acid synthetase messenger RNA

Fatty acid synthetase messenger RNA was obtained from rat liver polysomal RNA and then injected into Xenopus laevis oocytes. The radioactive fatty acid synthetase protein synthesized in the oocytes was identified by immunoprecipitation with anti-fatty acid synthetase antibody and the immunoprecipitate was then characterized by electrophoresis on sodium dodecyl sulfate-polyacrylamide gel. Co-migration of authentic fatty acid synthetase and the labeled product synthesized in oocytes was observed. Based on sucrose density gradient analysis, the rat liver fatty acid synthetase mRNA has a sedimentation coefficient of approximately 33 S, which agrees with the predicted minimum size necessary to code for the fatty acid synthetase protein. In addition, this mRNA was partially purified with oligo(dT)-cellulose, which indicates that it has a polyadenylate region. The relative in vivo rate of synthesis of fatty acid synthetase and the level of fatty acid synthetase mRNA in liver were also determined during the course of dietary induction of this enzyme. The results indicate that the dietary-induced increase in the level of fatty acid synthetase is probably due to an increased level of the fatty acid synthetase mRNA.     

Genetic enhancement of fatty acid synthesis by targeting rat liver ATP:citrate lyase into plastids of tobacco

ATP:citrate lyase (ACL) catalyzes the conversion of citrate to acetyl-coenzyme A (CoA) and oxaloacetate and is a key enzyme for lipid accumulation in mammals and oleaginous yeasts and fungi. To investigate whether heterologous ACL genes can be targeted and imported into the plastids of plants, a gene encoding a fusion protein of the rat liver ACL with the transit peptide for the small subunit of ribulose bisphosphate carboxylase was constructed and introduced into the genome of tobacco. This was sufficient to provide import of the heterologous protein into the plastids. In vitro assays of ACL in isolated plastids showed that the enzyme was active and synthesized acetyl-CoA. Overexpression of the rat ACL gene led to up to a 4-fold increase in the total ACL activity; this increased the amount of fatty acids by 16% but did not cause any major change in the fatty acid profile. Therefore, increasing the availability of acetyl-CoA as a substrate for acetyl-CoA carboxylase and subsequent reactions of fatty acid synthetase has a slightly beneficial effect on the overall rate of lipid synthesis in plants.     

Long-term regulation by theophylline of fatty acid synthetase, acetyl-CoA carboxylase and lipid synthesis in cultured glial cells.

The long-term regulation of fatty acid synthetase and acetyl-CoA carboxylase and of fatty acid and sterol synthesis was studied in C-6 glial cells in culture. When theophylline (10(-3) M) was added to the culture medium of these cells, rates of lipid synthesis from acetate and activities of synthetase and carboxylase became distinctly lower than in cells that were untreated. This effect appeared after approximately 12 h, and after 48 h enzymatic activities were reduced approx. 2-fold and rates of lipid synthesis from acetate 3- to 4-fold. The likelihood that the decrease in fatty acid synthesis from acetate was caused by the decrease in activities of fatty acid synthetase and acetyl-CoA carboxylase was established by several observations. These indicated that the locus of the effect probably did not reside at the level of acetate uptake into the cell, alterations in acetate pool sizes or conversion of acetate to acetyl-CoA. Moreover, de novo fatty acid synthesis was found to be the predominant pathway in these glial cells, whether treated with theophylline or not. The mechanism of the effect of theophylline on fatty acid synthetase was shown by immunochemical techniques to involve an alteration in content of enzyme rather than in catalytic efficiency. The change in content of fatty acid synthetase was shown by isotopic-immunochemical experiments to involve a decrease in synthesis of the enzyme. The mechanism whereby theophylline leads to a decrease in lipogenesis and in the synthesis of fatty acid synthetase may not be mediated entirely by inhibition of phosphodiesterase and an increase in cyclic AMP levels, because dibutyryl cyclic AMP (10(-3) M) only partially reproduced the effect.     

Molecular cloning of cDNA for rat mitochondrial 3-oxoacyl-CoA thiolase

Messenger RNA of rat 3-oxoacyl-CoA thiolase (acetyl-CoA acyltransferase), a mitochondrial matrix enzyme involved in fatty acid beta-oxidation, was enriched by immunoprecipitation of rat liver free polysomes and recombinant plasmids were prepared from the enriched mRNA by a modification of the vector-primer method of Okayama and Berg. The transformants were initially screened for 3-oxoacyl-CoA thiolase cDNA sequences by differential colony hybridization with [32P]cDNAs, synthesized from the immunopurified and unpurified mRNAs. The cDNA clones for 3-oxoacyl-CoA thiolase were identified by hybrid-arrested translation and hybrid-selected translation. One of the clones, designated pT1-1, contained a 700-base insert and hybridized to a mRNA species of 1.6 X 10(3) bases in rat liver. The transformants were rescreened using the cDNA insert of pT1-1 as a hybridization probe and a clone (pT1-19) with a 1.5 X 10(3)-base insert was obtained. Activity and concentration of 3-oxoacyl-CoA thiolase mRNA were quantified by in vitro translation and dot-blot analysis using the cDNA insert as a hybridization probe. The level of translatable and hybridizable mRNA in rat liver was increased about 5.1-fold and 4.6-fold, respectively, after administration of di-(2-ethylhexyl)phthalate, a potent inducer of the enzyme. The 3-oxoacyl-CoA thiolase mRNA levels thus determined correlated closely with levels of the activity and amount of this enzyme.     

Mammalian fatty acid synthetase. III. Characterization of human liver synthetase products and kinetics of methylmalonyl-CoA inhibition.

When propionyl-CoA was substituted for either acetyl-CoA or butyryl-CoA in the presence of [14C]malonyl-CoA and NADPH, the pure human liver fatty acids synthetase complex synthesized only straight-chain, saturated, 15- and 17-carbon radioactive fatty acids. At optimal concentrations, propionyl-CoA was a better primer of fatty acid synthesis than acetyl-CoA. Methylmalonyl-CoA inhibited the synthetase competitively with respect to malonyl-CoA. The Ki was calculated to be 8.4 muM. These findings provide an in vitro model and offer a direct explanation at the molecular level for some of the abnormal manifestations observed in diseases characterized by increased cellular concentrations of propionyl-CoA and methylmalonyl-CoA.     

Role of protein synthesis in the carbohydrate-induced changes in the activities of acetyl-CoA carboxylase and hydroxymethylglutaryl-CoA reductase in cultured rat hepatocytes.

Changes in the activities of acetyl-CoA carboxylase and HMG-CoA (3-hydroxy-3-methylglutaryl-CoA) reductase were studied in primary cultures of adult-rat hepatocytes after exposure of the cells to insulin and/or carbohydrates. To determine the contribution of protein synthesis to changes in enzyme activity, the relative rate of synthesis of each enzyme was measured and the amount of translatable mRNA coding for the enzymes was determined by translation in vitro and immunoprecipitation. Addition of insulin to the culture medium increased the activities of acetyl-CoA carboxylase and HMG-CoA reductase by approx. 4- and 3-fold respectively. Although similar increases in the relative rate of synthesis of each protein and template activity were noted, initial increases in the activity of each enzyme occurred before any changes in protein synthesis were observed, suggesting the involvement of post-translational modification of enzyme activity in addition to changes in protein synthesis. The addition of fructose to the culture medium, in the absence of insulin, increased the activity of the carboxylase and the reductase approx. 3-fold, similar to the effects of insulin. However, the effect of fructose was to increase the rate of synthesis and the amount of translatable mRNA coding for acetyl-CoA carboxylase, whereas the increase in the activity of HMG-CoA reductase was not accompanied by any changes in the rate of synthesis or template activity. The effects of fructose could not be mimicked by glucose unless insulin was also present in the culture medium. Similar to observations in vitro, the injection of insulin or the feeding of a high-fructose diet to rats made diabetic by the injection of streptozotocin produced an increase in the activities of acetyl-CoA carboxylase and HMG-CoA reductase, and only the increase in the activity of the carboxylase was accompanied by an increase in the amount of translatable mRNA coding for the enzyme. The results are discussed in terms of the effects of fructose on the synthesis of enzymes involved in lipogenesis.     

Regulation of fatty acid synthesis.

Acetyl-CoA carboxylase and fatty acid synthetase are the two major enzymes involved in the synthesis of fatty acids in animals. The activities of both enzymes are affected by nutritional manipulations. Although acetyl-CoA carboxylase is considered generally to be the rate-limiting step in lipogenesis, there is evidence that suggests that fatty acid synthetase may become rate limiting under certain conditions. The principal support for the view that acetyl-CoA carboxylase is the rate-limiting enzyme for lipogenesis is that the activity of the enzyme is controlled by allosteric effectors that change the catalytic efficiency of the enzyme. Until recently, the only known control of fatty acid synthetase was through changes in rate of enzyme synthesis. Data are reviewed that show that fatty acid synthetase can exist in forms possessing different catalytic activities. Thus fatty acid synthetase appears to be subject to the type of control necessary for an enzyme to serve as a regulator of the rate of a biological process over a short term.     

Estrogen regulation of hepatic 3-hydroxy-3-methylglutaryl coenzyme A reductase and acetyl-CoA carboxylase in xenopus laevis

Administration of estradiol-17 beta to male Xenopus laevis evokes the proliferation of the endoplasmic reticulum and the Golgi apparatus and the synthesis and secretion by the liver of massive amounts of the egg yolk precursor phospholipoglycoprotein, vitellogenin. We have investigated the effects of estrogen on three key regulatory enzymes in lipid biosynthesis, 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase, the major regulatory enzyme in cholesterol and isoprenoid synthesis, and acetyl-CoA carboxylase and fatty acid synthetase, which regulate fatty acid biosynthesis. HMG-CoA reductase activity and cholesterol synthesis increase in parallel following estrogen administration. Reductase activity in estrogen stimulated Xenopus liver cells peaks at 40-100 times the activity observed in control liver cells. The increased rate of reduction of HMG-CoA to mevalonic acid is not due to activation of pre-existing HMG-CoA reductase by dephosphorylation, as the fold induction is unchanged when reductase from control and estrogen-stimulated animals is fully activated prior to assay. The estrogen-induced increase of fatty acid synthesis is paralleled by a 16- to 20-fold increase of acetyl-CoA carboxylase activity, indicating that estrogen regulates fatty acid synthesis at the level of acetyl-CoA carboxylase. Fatty acid synthetase activity was unchanged during the induction of fatty acid biosynthesis by estrogen. The induction of HMG-CoA reductase and of acetyl-CoA carboxylase by estradiol-17 beta provides a useful model for regulation of these enzymes by steroid hormones.     

Mechanisms by which tumor necrosis factor stimulates hepatic fatty acid synthesis in vivo

We have previously shown that bolus intravenous administration of tumor necrosis factor (TNF) to normal rats results in a rapid (within 90 min) stimulation of hepatic fatty acid synthesis, which is sustained for 17 hr. We now demonstrate that TNF stimulates fatty acid synthesis by several mechanisms. Fatty acid synthetase and acetyl-CoA carboxylase (measured after maximal stimulation by citrate) were not higher in livers from animals that had been treated with TNF 90 min before study compared to controls. In contrast, 16 hr after treatment with TNF, fatty acid synthetase was slightly elevated (35%) while acetyl-CoA carboxylase was increased by 58%. To explain the early rise in the hepatic synthesis of fatty acids, we examined the regulation of acetyl-CoA carboxylase. The acute increase in fatty acid synthesis was not due to activation of acetyl-CoA carboxylase by change in its phosphorylation state (as calculated by the ratio of activity in the absence and presence of 2 mM citrate). However, hepatic levels of citrate, an allosteric activator of acetyl-CoA carboxylase, were significantly elevated (51%) within 90 min of TNF treatment. TNF also induces an acute increase (within 90 min) in the plasma levels of free fatty acids. However, hepatic levels of fatty acyl-CoA, which can inhibit acetyl-CoA carboxylase, did not rise 90 min following TNF treatment and were 35% lower than in control livers by 16 hr after TNF. These data suggest that TNF acutely regulates hepatic fatty acid synthesis in vivo by raising hepatic levels of citrate.(ABSTRACT TRUNCATED AT 250 WORDS)     

Molecular cloning of cDNA for acetyl-coenzyme A carboxylase

Poly(A)+ RNA from lactating rat mammary glands was size-fractionated to enrich the relative amount of acetyl-CoA carboxylase mRNA. The enriched mRNA was used to generate a lambda gt11 cDNA library. Initial screening with polyclonal antiserum to acetyl-CoA carboxylase produced three positive clones. Western blot analysis revealed that two clones, lambda DH3 and lambda KH18, synthesized 165,000-dalton proteins that were recognized by antibodies to acetyl-CoA carboxylase and beta-galactosidase, indicating that acetyl-CoA carboxylase/beta-galactosidase fusion proteins were produced. Competition experiments with purified acetyl-CoA carboxylase further demonstrated that the fusion proteins contained acetyl-CoA carboxylase protein segments. Antibodies which are specific to the fusion proteins were isolated. These antibodies cross-reacted only with acetyl-CoA carboxylase in a preparation of partially purified acetyl-CoA carboxylase. In addition, the antibodies immunoprecipitated enzyme activity from a crude liver homogenate. Northern blot analysis of total RNA revealed two RNA species: one 10 kilobases and the other 3.0 kilobases. The levels of these RNA species increased when starved animals were fed a fat-free diet, indicating that they are coordinately regulated.     

Effects of dietary nutrients on substrate and effector levels of lipogenic enzymes, and lipogenesis from tritiated water in rat liver

When fasted rats were refed for 4 days with a carbohydrate and protein diet, a carbohydrate diet (without protein) or a protein diet (without carbohydrate), the effects of dietary nutrients on the fatty acid synthesis from injected tritiated water, the substrate and effector levels of lipogenic enzymes and the enzyme activities were compared in the livers. In the carbohydrate diet group, although acetyl-CoA carboxylase was much induced and citrate was much increased, the activity of acetyl-CoA carboxylase extracted with phosphatase inhibitor and activated with 0.5 mM citrate was low in comparison to the carbohydrate and protein diet group. The physiological activity of acetyl-CoA carboxylase seems to be low. In the protein diet group, the concentrations of glucose 6-phosphate, acetyl-CoA and malonyl-CoA were markedly higher than in the carbohydrate and protein group, whereas the concentrations of oxaloacetate and citrate were lower. The levels of hepatic cAMP and plasma glucagon were high. The activities of acetyl-CoA carboxylase and also fatty acid synthetase were low in the protein group. By feeding fat, the citrate level was not decreased as much as the lipogenic enzyme inductions. Comparing the substrate and effector levels with the Km and Ka values, the activities of acetyl-CoA carboxylase and fatty acid synthetase could be limited by the levels. The fatty acid synthesis from tritiated water corresponded more closely to the acetyl-CoA carboxylase activity (activated 0.5 mM citrate) than to other lipogenic enzyme activities. On the other hand, neither the activities of glucose-6-phosphate dehydrogenase and malic enzyme (even though markedly lowered by diet) nor the levels of their substrates appeared to limit fatty acid synthesis of any of the dietary groups. Thus, it is suggested that under the dietary nutrient manipulation, acetyl-CoA carboxylase activity would be the first candidate of the rate-limiting factor for fatty acid synthesis with the regulations of the enzyme quantity, the substrate and effector levels and the enzyme modification.