Cerulenin markedly inhibited the growth of Acholeplasma laidlawii. A. axanthum and A. granularum were less susceptible, whereas the sterol-requiring Mycoplasma species examined showed very little susceptibility. The inhibition was not reversed by the addition of long-chain fatty acids to the medium. At a concentration of 20 μg/ml, cerulenin inhibited the incorporation of [14C]acetate into A. laidlawii membrane lipids, but it had no effect on either protein or nucleic acid biosynthesis. Cerulenin inhibited both the de novo synthesis of long-chain fatty acids and the elongation of medium-chain fatty acids. As a result, carotenoid biosynthesis was stimulated, and increased amounts of oleic and elaidic acids were incorporated into membrane polar lipids. Our studies support the concept that cerulenin can serve as a useful tool for obtaining better control of fatty acid composition of A. laidlawii membranes.
The antibiotic cerulenin markedly inhibits the growth of Escherichia coli. The effects of the antibiotic on cellular syntheses were studied by measuring the incorporation of labeled precursors into lipids and macromolecules. During the first 40 min after the addition of cerulenin to a culture of growing cells, lipid synthesis was inhibited more than 90% and ribonucleic acid and deoxyribonucleic acid synthesis about 25%, whereas protein synthesis was not affected. At later periods after cerulenin addition (1 to 2 h), the inhibition of cell growth and of lipid and protein synthesis was complete. Upon removal of cerulenin from the culture, growth was restored and lipid synthesis resumed more rapidly than did the synthesis of protein.
Addition of both palmitate and oleate, but not of either fatty acid alone, reversed the inhibition of growth by cerulenin. These findings support the conclusion that the antibiotic effects of cerulenin are due to a specific inhibition of fatty acid synthesis.
Germination of spores of the fungus Botryodiplodia theobromae was inhibited by the antilipogenic antibiotic cerulenin. The spores remained viable in the presence of the antibiotic, however, and after prolonged incubation they were able to overcome the inhibition. Cerulenin inhibition of germination was reversed by Tween 40 and Tween 60 (derivatives of palmitate and stearate, respectively), but not by representatives of a range of free fatty acids or their soaps. Cerulenin abolished incorporation of [14C]acetate into sterols and triglycerides and reduced its incorporation into fatty acids by 69%. Cyanide-sensitive oxygen consumption by spores incubated in the presence of cerulenin was greatly reduced throughout germination, and the activity of cytochrome c oxidase was no more than 13% of the activity in untreated spores, even after prolonged incubation. However, low-temperature difference spectra of mitochondrial extracts showed that the cerulenin-treated spores accumulated a threefold excess of cytochrome a, whereas the cellular concentrations of cytochroms c and b were identical to those of untreated spores. Cerulenin treatment sharply reduced the rates of whole spore protein and RNA synthesis. Cerulenin had no effects upon mitochondrial morphology which could be discerned with an electron microscope.
The formation of penicillinase by cultures of Bacillus licheniformis was preferentially suppressed by cerulenin, an antibiotic known to specifically inhibit fatty acid synthesis in microorganisms. The effect was studied at cerulenin concentrations that had almost no effect on the rate of cell growth and overall protein synthesis, but that reduced the rate of [14C]acetate incorporation (by 50 to 70%), indicating partial inhibition of lipid synthesis. The levels of both the released enzyme (exopenicillinase) and its cell-bound precursor were reduced to the same extent (70% to 80%). Enzyme formation was gradually resumed after the removal of cerulenin or the addition of a mixture of fatty acids prepared from lipids extracted from B. licheniformis. Reversal was less effective as the time interval between treatment with cerulenin and addition of fatty acids increased. We conclude that de novo synthesis of fatty acids is required for the formation of both the membrane-bound and extracellular penicillinase. Suppression of the membrane-bound enzyme is a likely consequence of the altered membrane (decreased lipid-to-lipid ratio and increased density) seen in cerulenin-treated preparations. The corresponding suppression of exopenicillinase is consistent with the view that it is derived from the membrane-bound form. A mechanism linking the general class of exportable proteins to specific aspects of lipid synthesis is discussed.
Exogenously supplied long-chain fatty acids have been shown to markedly alleviate the inhibition of phototrophic growth of cultures of Rhodopseudomonas sphaeroides caused by the antibiotic cerulenin. Monounsaturated and polyunsaturated C18 fatty acids were most effective in relieving growth inhibition mediated by cerulenin. Medium supplementation with saturated fatty acids (C14 to C18) failed to influence the inhibitory effect of cerulenin. The addition of mixtures of unsaturated and saturated fatty acids to the growth medium did not enhance the growth of cerulenin-inhibited cultures above that obtained with individual unsaturated fatty acids as supplements. Resolution and fatty acid analysis of the extractable lipids of R. sphaeroides revealed that exogenously supplied fatty acids were directly incorporated into cellular phospholipids. Cells treated with cerulenin displayed an enrichment in their percentage of total saturated fatty acids irrespective of the presence of exogenous fatty acids. Cerulenin produced comparable inhibitions of the rates of both fatty acid and phospholipid synthesis and was further found to preferentially inhibit unsaturated fatty acid synthesis.
Inhibition of de novo fatty acid biosynthesis by the antibiotic cerulenin in Bacillus subtilis stopped de novo synthesis of neutral lipids and phospholipids. The bacteria ceased growing but remained completely viable. Addition of 12-methyltetradecanoic acid and palmitic acid to the culture medium of cerulenin-treated cells restored growth of the bacteria, albeit at a reduced rate. Although the de novo synthesis of all lipid components of the membrane was blocked, citrate-Mg2+ transport activity remained inducible, and induced cells did not lose this transport activity when treated with cerulenin. Shortly after the addition of cerulenin, the rate of ribonucleic acid synthesis dropped rapidly and was followed by a slower decrease in the rate of protein synthesis. The rate of deoxyribonucleic acid synthesis remained almost unaffected. The rapid decrease of ribonucleic acid synthesis in cerulenin-treated cells might be due to the inhibition of de novo fatty acid biosynthesis or it might be due to a secondary effect of cerulenin in B. subtilis cells.
The effect of the fatty acid synthesis inhibitor cerulenin on growth and dextransucrase (EC 220.127.116.11) production by Streptococcus mutans 6715 was analyzed. Growth was markedly inhibited by less than 1 microgram of the antibiotic per ml. Under conditions where cerulenin did not inhibit amino acid incorporation into protein but did block acetate metabolism into lipid, the production of extracellular dextransucrase was suppressed. Inhibition was not due to a direct effect of the antibiotic on the enzyme or the lack of release of enzyme from the bacterial cell surface. Gel column chromatography demonstrated that enzyme produced in the presence of cerulenin was highly aggregated, similar to the control enzyme. Although the addition of ysophosphatidylcholine to enzyme which had been synthesized in the presence of cerulenin stimulated glucan formation from sucrose, the increase was not greater than that produced with the control enzyme. The differential inhibition of dextransucrase production by cerulenin indicates that enzyme secretion requires the production of lipid and may reflect the mechanism by whch this enzyme is transported from the bacterial cell.
Sporulation of Saccharomyces cerevisiae G2-2 was inhibited by the antibiotic cerulenin which is known to be a specific inhibitor of fatty acid and sterol synthesis. This inhibition was reversed by various fatty acids, especially by oleic acid (C18:1) and pentadecanoic acid (C15:0). Ergosterol showed only slight reversibility of this inhibition. When cerulenin was added to the sporulation medium later than 12 h after the start of incubation, the marked inhibition disappeared. When the fatty acid fraction extracted from the sporulated yeasts was added to the cells pretreated with cerulenin for more than 6 h, sporulation became evident 6 h after the fatty acid fraction addition. Therefore, sufficient synthesis of fatty acids required for sporulation was assumed to occur during the first 6 h in phase I of yeast sporulation.
The effects of growth temperature on the fatty acid composition of the phospholipids of the fungus Talaromyces thermophilus were investigated. This thermophilic organism was unable to increase the degree of unsaturation of its fatty acids when shifted from high to low growth temperatures. Inhibition of fatty acid synthesis by the antibiotic cerulenin was reversed by the addition of a mixture of palmitic, stearic, and oleic acids and ergosterol. The data obtained were consistent with the hypothesis that the thermophilic character of T. thermophilus is due to metabolic limitations that restrict its ability to regulate membrane fluidity.
When bakers' yeast cells were grown anaerobically in a medium supplemented with Tween 80 and ergosterol, exposure during aeration to the fatty acid synthesis inhibitor, cerulenin, had little effect upon respiratory adaptation, the induction of enzymes of electron transport, or the in vivo incorporation of [14C]leucine into mitochondrial membranes. These lipid-supplemented cells were apparently able to undergo normal respiratory adaptation utilizing endogenous lipids alone. The level of cerulenin used (2 μg/ml) inhibited the in vivo incorporation of [14C]acetate into mitochondrial membrane lipids by 96%. If, however, the cells were deprived of exogenous lipid during anaerobic growth, subsequent exposure to cerulenin severely reduced their capacity to undergo respiratory adaptation, to form enzymes of electron transport, and to incorporate amino acid into both total cell and mitochondrial membrane proteins. This cerulenin-mediated inhibition of enzyme formation and of protein synthesis was nearly completely reversed by the addition of exogenous lipid during the aeration of the cells. In lipid-limited cells, chloramphenicol also had dramatic inhibitory effects, both alone (75%) and together with cerulenin (85%), upon total cell and mitochondrial membrane [14C]leucine incorporation. This marked chloramphenicol-mediated inhibition was also largely reversed by exogenous lipid. It is concluded that, in lipid-limited cells, either cerulenin or chloramphenicol may prevent the emergence of a pattern of lipids required for normal levels of protein synthetic activity. The effect of cerulenin upon the formation of mitochondrial inner membrane enzymes thus appears to reflect a nonspecific effect of this antilipogenic antibiotic upon total cell protein synthesis.
This study investigated the biological significance of the inhibition of fatty acid synthase (FAS) in multiple myeloma (MM) using the small molecule inhibitor Cerulenin. Cerulenin triggered growth inhibition in both MM cell lines and MM patient cells, and overcame the survival and growth advantages conferred by interleukin-6, insulin-like growth factor-1, and bone marrow stromal cells. It induced apoptosis in MM cell lines with only modest activation of caspase -8, -9, -3 and PARP; moreover, the pan-caspase inhibitor Z-VAD-FMK did not inhibit Cerulenin-induced apoptosis and cell death. In addition, treatment of MM cells with Cerulenin primarily up-regulated apoptosis-inducing factor/endonuclease G, mediators of caspase-independent apoptosis. Importantly, Cerulenin induced endoplasmic reticulum stress response via up-regulation of the Grp78/IRE1α/JNK pathway. Although the C-Jun-NH2-terminal kinase (JNK) inhibitor SP600215 blocked Cerulenin-induced cytotoxicity, it did not inhibit apoptosis and caspase cleavage. Furthermore, Cerulenin showed synergistic cytotoxic effects with various agents including Bortezomib, Melphalan and Doxorubicin. Our results therefore indicate that inhibition of FAS by Cerulenin primarily triggered caspase-independent apoptosis and JNK-dependent cytotoxicity in MM cells. This report demonstrated that inhibition of FAS has anti-tumour activity against MM cells, suggesting that it represents a novel therapeutic target in MM.
multiple myeloma; fatty acid synthase; apoptosis; JNK
Bacterial bioluminescence is very sensitive to cerulenin, a fungal antibiotic which is known to inhibit fatty acid synthesis. When Vibrio harveyi cells pretreated with cerulenin were incubated with [3H]myristic acid in vivo, acylation of the 57-kilodalton reductase subunit of the luminescence-specific fatty acid reductase complex was specifically inhibited. In contrast, in vitro acylation of both the synthetase and transferase subunits, as well as the activities of luciferase, transferase, and aldehyde dehydrogenase, were not adversely affected by cerulenin. Light emission of wild-type V. harveyi was 20-fold less sensitive to cerulenin at low concentrations (10 micrograms/ml) than that of the dark mutant strain M17, which requires exogenous myristic acid for luminescence because of a defective transferase subunit. The sensitivity of myristic acid-stimulated luminescence in the mutant strain M17 exceeded that of phospholipid synthesis from [14C]acetate, whereas uptake and incorporation of exogenous [14C]myristic acid into phospholipids was increased by cerulenin. The reductase subunit could be labeled by incubating M17 cells with [3H]tetrahydrocerulenin; this labeling was prevented by preincubation with either unlabeled cerulenin or myristic acid. Labeling of the reductase subunit with [3H]tetrahydrocerulenin was also noted in an aldehyde-stimulated mutant (A16) but not in wild-type cells or in another aldehyde-stimulated mutant (M42) in which [3H]myristoyl turnover at the reductase subunit was found to be defective. These results indicate that (i) cerulenin specifically and covalently inhibits the reductase component of aldehyde synthesis, (ii) this enzyme is partially protected from cerulenin inhibition in the wild-type strain in vivo, and (iii) two dark mutants which exhibit similar luminescence phenotypes (mutants A16 and M42) are blocked at different stages of fatty acid reduction.
Lipids are a source of metabolic energy, as well as essential components of cellular membranes. Although they have been shown to be key players in the regulation of cell proliferation in various eukaryotes, including microalgae, their role in the cell cycle of cnidarian-dinoflagellate (genus Symbiodinium) endosymbioses remains to be elucidated. The present study examined the effects of a lipid synthesis inhibitor, cerulenin, on the cell cycle of both cultured Symbiodinium (clade B) and those engaged in an endosymbiotic association with the sea anemone Aiptasia pulchella. In the former, cerulenin exposure was found to inhibit free fatty acid (FFA) synthesis, as it does in other organisms. Additionally, while it also significantly inhibited the synthesis of phosphatidylethanolamine (PE), it did not affect the production of sterol ester (SE) or phosphatidylcholine (PC). Interestingly, cerulenin also significantly retarded cell division by arresting the cell cycles at the G0/G1 phase. Cerulenin-treated Symbiodinium were found to be taken up by anemone hosts at a significantly depressed quantity in comparison with control Symbiodinium. Furthermore, the uptake of cerulenin-treated Symbiodinium in host tentacles occurred much more slowly than in untreated controls. These results indicate that FFA and PE may play critical roles in the recognition, proliferation, and ultimately the success of endosymbiosis with anemones.
The antibiotic cerulenin differentially inhibited the production of glucosyltransferase activity by Streptococcus mutans GS5. Cerulenin preferentially inhibited [14C]acetate incorporation into cellular lipids but did not affect protein synthesis or ribonucleic acid synthesis in the same manner. No significant intracellular accumulation of glucosyltransferase activity could be demonstrated in cultures treated with cerulenin. On the other hand, another inhibitor of lipid synthesis, sodium chlorophenoxyisobutyrate, did not differentially inhibit glucosyltransferase expression. In addition, the role of a metal-requiring protease in the production of glucosyltransferase activity was suggested by the observation that the chelator quinacrine differentially inhibited the production of the enzyme.
There is considerable evidence correlating the production of increased proportions of membrane unsaturated fatty acids (UFAs) with bacterial growth at low temperatures or high pressures. In order to assess the importance of UFAs to microbial growth under these conditions, the effects of conditions altering UFA levels in the psychrotolerant piezophilic deep-sea bacterium Photobacterium profundum SS9 were investigated. The fatty acids produced by P. profundum SS9 grown at various temperatures and pressures were characterized, and differences in fatty acid composition as a function of phase growth, and between inner and outer membranes, were noted. P. profundum SS9 was found to exhibit enhanced proportions of both monounsaturated (MUFAs) and polyunsaturated (PUFAs) fatty acids when grown at a decreased temperature or elevated pressure. Treatment of cells with cerulenin inhibited MUFA but not PUFA synthesis and led to a decreased growth rate and yield at low temperature and high pressure. In addition, oleic acid-auxotrophic mutants were isolated. One of these mutants, strain EA3, was deficient in the production of MUFAs and was both low-temperature sensitive and high-pressure sensitive in the absence of exogenous 18:1 fatty acid. Another mutant, strain EA2, produced little MUFA but elevated levels of the PUFA species eicosapentaenoic acid (EPA; 20:5n-3). This mutant grew slowly but was not low-temperature sensitive or high-pressure sensitive. Finally, reverse genetics was employed to construct a mutant unable to produce EPA. This mutant, strain EA10, was also not low-temperature sensitive or high-pressure sensitive. The significance of these results to the understanding of the role of UFAs in growth under low-temperature or high-pressure conditions is discussed.
The viability of Streptococcus lactis and Lactobacillus sp. A-12 after freezing at -17°C for 48 h was better preserved when the cells were grown in medium supplemented with oleic acid or Tween 80 (polyoxyethylene sorbitan monooleate). A pronounced change in the cellular fatty acid composition was noted when the bacteria were grown in the presence of Tween 80. In S. lactis the ratio of unsaturated to saturated fatty acids increased from 1.18 to 2.55 and in Lactobacillus sp. A-12 it increased from 0.85 to 1.67 when Tween 80 was added to the growth medium. The antibiotic cerulenin markedly inhibited the growth of lactic acid bacteria in tomato juice (TJ) medium but had almost no effect on the growth of the bacteria in TJ medium containing Tween 80 (or oleic acid). The antibiotic inhibited markedly the incorporation of [1-14C]acetate but had no inhibitory effect on the incorporation of exogenous [1-14C]oleate (or [1-14C]palmitate) into the lipid fractions of lactic acid bacteria. Thus, the fatty acid composition of lactic acid bacteria, inhibited by the antibiotic cerulenin, can be modulated by exogenously added oleic acid (or Tween 80) without the concurrent endogenous fatty acid synthesis from acetate. The data obtained suggest that cerulenin inhibits neither cyclopropane fatty acid synthesis nor elongation of fatty acid acyl intermediates. The radioactivity of cells grown in the presence of [1-14C]oleate and cerulenin was associated mainly with cyclopropane Δ19:0, 20:0 + 20:1, and 21:0 acids. As a consequence, cerulenin caused a decrease in the ratio of unsaturated to saturated fatty acids in lactic acid bacteria as compared with cells grown in TJ medium plus Tween 80 but without cerulenin. Cerulenin caused a decrease in the viability of S. lactis and Lactobacillus sp. A-12 after freezing at -17°C for 48 h only when Tween 80 was present in the growth medium. We conclude that the sensitivity of lactic acid bacteria to damage from freezing can be correlated with specific alterations in the cellular fatty acids.
CM-55 is a synthetic analogue of the antibiotic cerulenin with the chemical structure of 2, 3-dodecenyl-4-oxo-dimethyl amide. This compound inhibited the growth of Saccharomyces cerevisiae ATCC 12341 and inhibited protein and lipid synthesis by 91 and 95%, respectively, at a concentration of 50 μg/ml (2.1 × 10−4 M). The inhibition of protein synthesis was associated with the partial reduction of ribonucleic acid synthesis and leucine transport. The mechanism of inhibition of lipid synthesis was further investigated in a cell-free extract of the yeast. CM-55 inhibited the incorporation of [14C]acetyl Coenzyme A (CoA) into both fatty acid (FAF) and non-saponifiable fractions (NSF). However, it did not inhibit [14C]malonyl CoA incorporation into FAF and only slightly inhibited [14C]mevalonate incorporation into NSF. The activity of 3-hydroxy-3-methylglutaryl CoA (HMG-CoA) synthase was inhibited more strongly than the incorporation of [14C]3-hydroxy-3-methylglutaryl CoA into NSF; this could account for the CM-55 inhibition of [14C]acetyl CoA incorporation into NSF.
Extracellular fructosyltransferase (levansucrase; EC 18.104.22.168) production in Actinomyces viscosus T14AV was demonstrated to occur concomitantly with cellular growth. The inhibition of both cellular ribonucleic acid and protein synthesis resulted in no further accumulation of enzyme activity. The antibiotic sodium clofibrate differentially inhibited the production of fructosyltransferase by strain T14AV. Furthermore, the antibiotic preferentially inhibited [14C]acetate incorporation into cellular lipid, but did not affect protein synthesis. In addition, no inhibition of fructosyltransferase production was observed upon the addition of the fatty acid acid synthesis inhibitor cerulenin. On the other hand, extracellular fructosyltransferase production was apparently stimulated in the presence of the cell wall synthesis inhibitors penicillin, amphomycin, and tunicamycin. These results are discussed in terms of the mechanism of extracellular protein production in A. viscosus.
Cerulenin, an inhibitor of fatty acid biosynthesis, has been used to study the role of the plasma membrane in germination of Candida albicans. To further elucidate this association, spontaneous, cerulenin-resistant mutants of C. albicans were isolated. Two of the mutants, 4918-2 and 4918-10, were compared biochemically with wild-type cells (4918). All strains grew equally well at 37 degrees C and synthesized fatty acids at comparable rates in the absence of the drug. In the presence of cerulenin, wild-type cells did not proceed through a logarithmic growth stage and exhibited a significantly impaired ability to incorporate [3H]acetate into newly synthesized lipid material. All strains were examined ultrastructurally. Alterations were observed in the membranous structures of cerulenin-treated wild-type cells. Such changes were not observed in cerulenin-treated mutant strains. Further examination of mutant strains revealed differences in cell wall protein and polysaccharide compositions when compared with those of wild-type cells. These apparent alterations in cell surface components may be correlated with the reduced abilities of mutant strains to adhere, in vitro, to mammalian cells.
Acholeplasma laidlawii was grown with different fatty acids for membrane lipid synthesis (saturated straight- and branched-chain acids and mono- and di-unsaturated acids). The ability of 12 different sterols to affect cell growth, lipid head group composition, the order parameter of the acyl chains, and the phase equilibria of in vivo lipid mixtures was studied. The following two effects were observed with respect to cell growth: with a given acyl chain composition of the membrane lipids, growth was stimulated, unaffected, reduced, or completely inhibited (lysis), depending on the sterol structure; and the effect of a certain sterol depended on the acyl chain composition (most striking for epicoprostanol, cholest-4-en-3-one, and cholest-5-en-3-one, which stimulated growth with saturated acyl chains but caused lysis with unsaturated chains). The three lytic sterols were the only sterols that caused a marked decrease in the ratio between the major lipids monoglucosyldiglyceride and diglucosyldiglyceride and hence a decrease in bilayer stability when the membranes were enriched in saturated (palmitoyl) chains. With these chains correlations were found for several sterols between the glucolipid ratio and the order parameter of the acyl chains, as well as the lamellar-reversed hexagonal phase transition, in model systems. A shaft experiment revealed a marked decrease in the ratio of monoglucosyldiglyceride to diglucosyldiglyceride with the lytic sterols in unsaturated (oleoyl) membranes. The two cholestenes induced nonlamellar phases in in vivo mixtures of oleoyl A. laidlawii lipids. The order parameters of the oleoyl chains were almost unaffected by the sterols. Generally, the observed effects cannot be explained by an influence of the sterols on the gel-to-liquid crystalline phase transition.
Bacillus subtilis B secretes an inducible, extracellular enzyme, levansucrase. Inhibition studies were undertaken to investigate the possible mechanism of release of this enzyme. The antibiotic cerulenin, at a concentration of 10 micrograms/ml, totally inhibited de novo lipid synthesis in B. subtilis B for at least 1 h, while only slightly reducing protein and RNA synthesis. At this concentration cerulenin, added concomitantly with the inducer sucrose, prevented the release of levansucrase for at least 150 min. This was not due to the prevention of inducer uptake by the cells. The release of the enzyme was also independent of cell division. In B. subtilis 1007 the induction of beta-galactosidase by 5 mM lactose was not prevented by cerulenin. Preliminary evidence indicated the association of a lipid moiety with the enzyme as it passes through the cytoplasmic membrane. Quinacrine (0.2 mM), which inhibits the penicillinase-releasing protease of Bacillus licheniformis, inhibited levansucrase release from B. subtilis B, but had no effect on lipid synthesis.
In the presence of an inhibitory concentration of cerulenin, cells of Staphylococcus aureus can resume growth when supplemented with either a saturated or an unsaturated fatty acid. A requirement for both types of acids for growth could not be demonstrated.
Liposarcomas (LS) are mesenchymal tumors that can recur after surgical resection and often do not respond to presently available medical therapies. This study demonstrates the dependence of LS on de novo long-chain fatty acid synthesis for growth. Lipogenesis can be impaired by inhibiting the activities of lipogenic enzymes, including acetyl CoA-carboxylase (ACC) and fatty acid synthase (FASN), or by suppressing the expression of key genes involved in the pathway and its regulation. The FASN inhibitors cerulenin and orlistat reduced the growth of two LS cell lines (LiSa2, SW872), as did inhibition of ACC with soraphen A. CDDO-Me, a synthetic triterpenoid, suppressed expression of Spot 14 and FASN genes and likewise inhibited LS cell growth. Importantly, the anti-proliferative effect of each agent was prevented by the co-administration of palmitate, the major product of cellular long-chain fatty acid synthesis. In stark contrast to LS cells, these compounds had no effect on the growth of fibroblasts. Four biochemically distinct agents that target critical points in the fatty acid synthetic pathway exert anti-proliferative effects on LS cells, and rescue of cell growth by palmitic acid suggests that reduced tumor cell lipogenesis mediates the growth inhibition. These findings warrant further studies aimed at the clinical exploitation of the dependence of LS cell growth on fatty acids.
fatty acid synthase; liposarcoma; spot 14
Exponential-phase cells of Streptococcus faecalis ATCC 9790 were treated with a concentration of cerulenin (5 micrograms/ml) that has been shown to block both lipoteichoic acid and lipid synthesis and cell division within 10 min. The morphological effect of this treatment was studied by making three-dimensional reconstructions of cells based on measurements taken from axial thin sections. This analysis indicated that cerulenin interferes with cell division by inhibiting normal constriction of the division furrow and centripetal growth of the cross wall in envelope growth sites. Rather than dividing, many of the sites in treated cells apparently continue to elongate and produce abnormally large amounts of peripheral wall surface. These observations were interpreted in terms of a previously proposed model in which cerulenin would prevent the synthesis of a lipid-containing inhibitor of autolytic enzyme activity needed for division. In addition, measurements showed that the average number of envelope growth sites per cell increased during treatment, suggesting that although cerulenin treatment blocks division, it does not interfere with the formation of new envelope growth sites. It was also observed that the size and frequency of mesosomes did not decline during the 60-min period of drug treatment. This tends to decrease the likelihood that mesosomes are formed from a pool of intracellular membrane precursors that would be depleted during a period of restricted lipid biosynthesis.
Thiolactomycin (TLM) and cerulenin are antibiotics that block Escherichia coli growth by inhibiting fatty acid biosynthesis at the beta-ketoacyl-acyl carrier protein synthase I step. Both TLM and cerulenin trigger the accumulation of intracellular malonyl-coenzyme A coincident with growth inhibition, and the overexpression of synthase I protein confers resistance to both antibiotics. Strain CDM5 was derived as a TLM-resistant mutant but remained sensitive to cerulenin. TLM neither induced malonyl-coenzyme A accumulation nor blocked fatty acid production in vivo; however, the fatty acid synthase activity in extracts from strain CDM5 was sensitive to TLM inhibition. The TLM resistance gene in strain CDM5 was mapped to 57.5 min of the chromosome and was an allele of the emrB gene. Disruption of the emrB gene converted strain CDM5 to a TLM-sensitive strain, and the overexpression of the emrAB operon conferred TLM resistance to sensitive strains. Thus, activation of the emr efflux pump is the mechanism for TLM resistance in strain CDM5.