Propionate represses the dnaA gene via the methylcitrate pathway-regulating transcription factor, PrpR, in Mycobacterium tuberculosis

Abstract During infection of macrophages, Mycobacterium tuberculosis, the pathogen that causes tuberculosis, utilizes fatty acids as a major carbon source. However, little is known about the coordination of the central carbon metabolism of M. tuberculosis with its chromosomal replication, particularly during infection. A recently characterized transcription factor called PrpR is known to directly regulate the genes involved in fatty acid catabolism by M. tuberculosis. Here, we report for the first time that PrpR also regulates the dnaA gene, which encodes the DnaA initiator protein responsible for initiating chromosomal replication. Using cell-free systems and intact cells, we demonstrated an interaction between PrpR and the dnaA promoter region. Moreover, real-time quantitative reverse-transcription PCR analysis revealed that PrpR acts as a transcriptional repressor of dnaA when propionate (a product of odd-chain-length fatty acid catabolism) was used as the sole carbon source. We hypothesize that PrpR may be an important element of the complex regulatory system(s) required for tubercle bacilli to survive within macrophages, presumably coordinating the catabolism of host-derived fatty acids with chromosomal replication.

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Mycobacterial fatty acid catabolism is repressed by FdmR to sustain lipogenesis and virulence

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2019305118 ◽

2021 ◽

Vol 118 (16) ◽

pp. e2019305118

Author(s):

Wenyue Dong ◽

Xiaoqun Nie ◽

Hong Zhu ◽

Qingyun Liu ◽

Kunxiong Shi ◽

...

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Carbon Source ◽

Fatty Acid Synthase ◽

Cell Envelope ◽

Flux Analysis ◽

Type I ◽

Chain Acyl ◽

Fatty Acid Catabolism ◽

Long Chain

Host-derived fatty acids are an important carbon source for pathogenic mycobacteria during infection. How mycobacterial cells regulate the catabolism of fatty acids to serve the pathogenicity, however, remains unknown. Here, we identified a TetR-family transcriptional factor, FdmR, as the key regulator of fatty acid catabolism in the pathogen Mycobacterium marinum by combining use of transcriptomics, chromatin immunoprecipitation followed by sequencing, dynamic 13C-based flux analysis, metabolomics, and lipidomics. An M. marinum mutant deficient in FdmR was severely attenuated in zebrafish larvae and adult zebrafish. The mutant showed defective growth but high substrate consumption on fatty acids. FdmR was identified as a long-chain acyl-coenzyme A (acyl-CoA)–responsive repressor of genes involved in fatty acid degradation and modification. We demonstrated that FdmR functions as a valve to direct the flux of exogenously derived fatty acids away from β-oxidation toward lipid biosynthesis, thereby avoiding the overactive catabolism and accumulation of biologically toxic intermediates. Moreover, we found that FdmR suppresses degradation of long-chain acyl-CoAs endogenously synthesized through the type I fatty acid synthase. By modulating the supply of long-chain acyl-CoAs for lipogenesis, FdmR controls the abundance and chain length of virulence-associated lipids and mycolates and plays an important role in the impermeability of the cell envelope. These results reveal that despite the fact that host-derived fatty acids are used as an important carbon source, overactive catabolism of fatty acids is detrimental to mycobacterial cell growth and pathogenicity. This study thus presents FdmR as a potentially attractive target for chemotherapy.

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The ability of ectomycorrhizal fungi to utilize fatty acids and a lipid as a carbon source for mycelial growth

Canadian Journal of Botany ◽

10.1139/b03-111 ◽

2003 ◽

Vol 81 (12) ◽

pp. 1285-1292 ◽

Cited By ~ 2

Author(s):

Takefumi Hattori ◽

Akira Ohta ◽

Masayuki Itaya ◽

Mikio Shimada

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Carbon Source ◽

Ectomycorrhizal Fungi ◽

Mycelial Growth ◽

The Other ◽

Other Hand ◽

The Family ◽

Ecm Fungi ◽

Relative Utilization

We have investigated growth of ectomycorrhizal (ECM) fungi (i.e., 55 strains of 32 species in 15 genera) on saturated (palmitate), monounsaturated (oleate), diunsaturated (linoleate), triunsaturated (linolenate) fatty acids, and the triacylglyceride of oleate (triolein) lipid to elucidate an ability to utilize the fatty acids and lipid as a carbon source for growth. Relative utilization ratios (URs, %) based on mycelial growth on glucose suggest that ECM fungi belonging to the family Thelephoraceae have an ability to utilize palmitate. On the other hand, ECM fungi in the genus Laccaria can utilize at least either palmitate or oleate. Furthermore, Hygropharus russula grows on palmitate, oleate, and slightly on triolein. Lactarius chrysorrheus grows only on palmitate. These fatty-acid- and lipid-utilizing fungi may be promising as model fungi for further elucidation of the metabolic ability to utilize the fatty acids and lipid as a carbon source. On the contrary, the fungi in the genus Suillus were shown to scarcely utilize the fatty acids and lipid. Furthermore, most ECM fungi did not grow on either linoleate or linolenate.Key words: carbon source, ectomycorrhizal fungi, fatty acid, lipid, mycelial growth.

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The Outer Surface Lipoprotein VolA Mediates Utilization of Exogenous Lipids by Vibrio cholerae

mBio ◽

10.1128/mbio.00305-13 ◽

2013 ◽

Vol 4 (3) ◽

Cited By ~ 23

Author(s):

Aaron C. Pride ◽

Carmen M. Herrera ◽

Ziqiang Guan ◽

David K. Giles ◽

M. Stephen Trent

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Carbon Source ◽

Outer Membrane ◽

Vibrio Cholerae ◽

Homeoviscous Adaptation ◽

Gram Negative ◽

Content Type ◽

Enzymatic Function ◽

Gram Negative Organisms

ABSTRACTPrevious work from our laboratory showed that the Gram-negative aquatic pathogenVibrio choleraecan take up a much wider repertoire of fatty acids than other Gram-negative organisms. The current work elaborated on the ability ofV. choleraeto exploit an even more diverse pool of lipid nutrients from its environment. We have demonstrated that the bacterium can use lysophosphatidylcholine as a metabolite for growth. Using a combination of thin-layer chromatography and mass spectrometry, we also showed that lysophosphatidylcholine-derived fatty acid moieties can be used for remodeling theV. choleraemembrane architecture. Furthermore, we have identified a lysophospholipase, VolA (Vibrioouter membrane lysophospholipase A), required for these activities. The enzyme is well conserved inVibriospecies, is coexpressed with the outer membrane fatty acid transporter FadL, is one of very few surface-exposed lipoprotein enzymes to be identified in Gram-negative bacteria and the first instance of a surface lipoprotein phospholipase. We propose a model whereby the bacterium efficiently couples the liberation of fatty acid from lysophosphatidylcholine to its subsequent metabolic uptake. An expanded ability to scavenge diverse environmental lipids at the bacterial surface increases overall bacterial fitness and promotes homeoviscous adaptation through membrane remodeling.IMPORTANCEOur understanding of how bacteria utilize environmental lipid sources has been limited to lipids such as fatty acids and cholesterol. This narrow scope may be attributed to both the intricate nature of lipid uptake mechanisms and the diversity of lipid substrates encountered within an ecological niche. By examining the ability of the pathogenVibrio choleraeto utilize exogenous lipids, we uncovered a surface-exposed lipoprotein (VolA) that is required for processing the prevalent host lipid lysophosphatidylcholine. VolA functions as a lipase liberating a fatty acid from exogenous lysophospholipids. The freed fatty acid is then transported into the cell, serving as a carbon source, or shunted into phospholipid synthesis for membrane assembly. A limited number of surface-exposed lipoproteins have been found in Gram-negative organisms, and few have enzymatic function. This work highlights the ability of bacteria to exploit exogenous lipids for both maintenance of the membrane and carbon source acquisition.

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The Largest Open Reading Frame (pks12) in the Mycobacterium tuberculosis Genome Is Involved in Pathogenesis and Dimycocerosyl Phthiocerol Synthesis

Infection and Immunity ◽

10.1128/iai.71.7.3794-3801.2003 ◽

2003 ◽

Vol 71 (7) ◽

pp. 3794-3801 ◽

Cited By ~ 49

Author(s):

Tatiana D. Sirakova ◽

Vinod S. Dubey ◽

Hwa-Jung Kim ◽

Michael H. Cynamon ◽

Pappachan E. Kolattukudy

Keyword(s):

Fatty Acids ◽

Mycobacterium Tuberculosis ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Immunoblot Analysis ◽

Open Reading Frame ◽

Pcr Analysis ◽

Reading Frame ◽

Sds Page

ABSTRACT The cell wall lipids in Mycobacterium tuberculosis are probably involved in pathogenesis. The largest open reading frame in the genome of M. tuberculosis H37Rv, pks12, is unique in that it encodes two sets of domains needed to produce fatty acids. A pks12-disrupted mutant was produced, and disruption was confirmed by both PCR analysis and Southern blotting. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed that a 430-kDa protein band present in the wild type was missing in the mutant. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MS) and liquid chromatography (LC)-MS analysis of tryptic peptides showed that 54 peptides distributed throughout this protein matched the pks12-encoded sequence. Biochemical analysis using [1-14C]propionate as the radiotracer showed that the pks12 mutant was deficient in the synthesis of dimycocerosyl phthiocerol (DIM). SDS-PAGE, immunoblot analysis of proteins, and analysis of fatty acids showed that the mutant can produce mycocerosic acids. Thus, the pks12 gene is probably involved in the synthesis of phthiocerol, the diol required for DIM synthesis. Growth of the pks12 mutant was attenuated in mouse alveolar macrophage cell line MH-S, and the virulence of the mutant in vivo was highly attenuated in a murine model. Thus, pks12 probably participates in DIM production and its expression is involved in pathogenesis.

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The Trans-Fatty Acid, Elaidic Acid, Inhibits Macrophage Fatty Acid Catabolism and Stimulates Expression of Inflammatory Mediators

Blood ◽

10.1182/blood.v120.21.3277.3277 ◽

2012 ◽

Vol 120 (21) ◽

pp. 3277-3277

Author(s):

Lisa J Robinson ◽

Janelle Zacherl ◽

Harry C Blair ◽

Stephanie J Mihalik

Keyword(s):

Gene Expression ◽

Fatty Acids ◽

Oleic Acid ◽

Fatty Acid ◽

Stearic Acid ◽

Inflammatory Mediators ◽

Elaidic Acid ◽

Trans Fatty Acids ◽

Trans Fat ◽

Fatty Acid Catabolism

Abstract Abstract 3277 In recent decades, addition to the diet of synthetically hydrogenated vegetable oils has markedly increased human consumption of trans fatty acids. Epidemiological studies have linked this change in diet to current high rates of atherosclerotic cardiovascular disease. Despite recognition of this important connection, the basic mechanisms by which trans fatty acids contribute to the pathogenesis of atherosclerosis are still not well understood. In the present studies we examined the effects of trans fatty acids on macrophage functions and their possible role in the pathogenesis of atherosclerosis. Human macrophages, derived from peripheral blood mononuclear cells, were treated with the trans fat elaidic acid (C18:Δ9–10 trans), the corresponding cis fatty acid oleic acid (C18:Δ9–10 cis), or the saturated fatty acid stearic acid (C18:0). We examined changes in macrophage fat metabolism using GC/MS to measure cell fatty acid content and intermediates, and MS/MS to identify acylcarnitine derivatives, and assayed fatty acid oxidation using fatty acids radiolabeled at the [1–14C] position and the double bond at the [C9-C103H] position. After 44 hours treatment with 100 micromolar elaidic acid, macrophages showed an accumulation of multiple unsaturated fatty acid intermediates, both long-chain and short-chain, by GC/MS analysis, that were not observed in cultures containing either oleic or stearic acid. Using acylcarnitine analysis, we observed an increase in C12 and C18 intermediates in the macrophages exposed to trans fat (either as fatty acids or partially hydrogenated soy oil) compared to controls. These results suggest a block in acyl-CoA removal one group proximate to the trans bond. Beta-oxidation assays using carbon-1 radiolabeled oleic and elaidic acids revealed enhanced entry of the trans-fat into the catabolic cycle compared to the entry of the natural cis-fatty acid. Using carbon 9–10 radiolabeled oleic acid to study oleic acid catabolism, we discovered that in the presence of the trans fat, oxidation of the cis fat was diminished. Thus, in addition to the block in the catabolism of the trans fat itself, the degradation of the cis monounsaturated fatty acids are also impaired in the presence of the trans fat. We then examined the effects of inhibited fatty acid catabolism on macrophage function by examining changes in gene expression. Initial results from Affymetrix gene expression profiling, were confirmed using quantitative real time PCR. These studies revealed that exposure to trans fatty acid, compared to cis fatty acids, markedly upregulated macrophage expression of interleukin 1 beta, an inflammatory cytokine previously implicated in the pathogenesis of atherosclerosis. Also increased was expression of heparin-binding epidermal growth factor, previously implicated as a stimulus for vascular smooth muscle proliferation in atherosclerosis. The results overall suggest that the deleterious effects of trans fats may be linked to impaired macrophage fatty acid catabolism, contributing to lipid accumulation in the atheroma, and also to increased macrophage production of inflammatory mediators. Disclosures: No relevant conflicts of interest to declare.

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Bioengineering of emulsifier structure: emulsan analogs

Canadian Journal of Microbiology ◽

10.1139/m97-053 ◽

1997 ◽

Vol 43 (4) ◽

pp. 384-390 ◽

Cited By ~ 18

Author(s):

Alexander Gorkovenko ◽

Jinwen Zhang ◽

Richard A. Gross ◽

Alfred L. Allen ◽

David L. Kaplan

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Carbon Source ◽

Chain Length ◽

Acinetobacter Calcoaceticus ◽

Chain Structure ◽

Fatty Acid Esters ◽

Side Chain ◽

Rag 1 ◽

Chain Lengths

Strategies were investigated to modulate the side chain structure of emulsans formed by Acinetobacter calcoaceticus RAG-1. Analysis of emulsan fatty acid side chain groups by gas chromatography – mass spectrometry (GC–MS) revealed that by providing the exogenous n-alkanoic fatty acids 15:0, 16:0, and 17:0, emulsan analogs were formed with 53, 46, and 44 mol%, respectively, of fatty acid substituents with chain lengths equal to that of the carbon source. In contrast, the increase in emulsan fatty acids of chain lengths less than 15 or greater than 17 by providing corresponding shorter and longer chain length fatty acids as carbon sources was not substantial. When [1-13C]-labeled (99% enriched) palmitic acid was used as a carbon source along with acetate, analysis of m/z 75/14 and 87/88 isotopomer ratios by GC-MS indicated that 84 and 86% of the 16:0 and 16:1 (9-cis) side groups, respectively, were incorporated intact from the 16:0 carbon source. The percentage of 14-, 15-, 16-, 17-, and 18-carbon chain length fatty acid esters that were monounsaturated were 11, 26, 50, 70, and 85%, respectively. Based on the observed percentage of unsaturated chain length dependence and almost identical enrichment at C-1 of 16:0 and 16:1 (9-cis) side groups from [1-13C]-labeled experiments, it was concluded that desaturation of preformed n-alkanoic acids was the predominant mechanism of their formation. Further work established correlations between side chain structure and product emulsification specificity/activity, so that bioengineered emulsans with improved selectivity can now be formed.Key words: emulsan, Acinetobacter calcoaceticus RAG-1, fatty acids, direct incorporation, emulsification activity.

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Pengaruh sumber karbon terhadap produksi bioplastik polihidroksialkanoat (PHA) dengan ralstonia eutropha

Jurnal Teknik Kimia Indonesia ◽

10.5614/jtki.2010.9.1.4 ◽

2018 ◽

Vol 9 (1) ◽

pp. 28

Author(s):

Martha Aznury ◽

Tjandra Setiadi ◽

Adi Pancoro

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Carbon Source ◽

Fermentation Process ◽

Volatile Fatty Acids ◽

Ralstonia Eutropha ◽

Operating Conditions ◽

Batch Bioreactor ◽

Pha Production ◽

Fructose Fermentation

Bioplastic Polyhidroxyalknoate (PHA) is a polyester type bioplastic with physicochemical properties resemble to those of polypropilen from petroleum. PHA production was investigated to determine the effect of carbon source on the fermentation process by Ralstonia eutropha. Specifically, Ralstonia eutropha was cultivated in a batch bioreactor to show the dynamics of P(3HB-co-3HV) copolymer production from glucose or fructose as C source. In adition, the effect of volatile fatty acids addition, as stimulator to the copolymer production, was also studied. The operating conditions in a 7 L bioreactor were at temperature 30 oC and pH 7.0. The concentration of carbon source glucose or fructose was 40 g/L, and after 20 hour fermentation, volatile fatty acids were added. With volatile fatty acids addition, the resulting fructose fermentation had PHA content of 32.78%, in which the HV percentage was 11.78%. Meanwhile, the fermentation of glucose, stimulated by volatile fatty acids, gave PHA as much as 20.19% with HV percentage of 8.71%. Therefore,, the Ralstonia eutropha fermentation of fructose as the carbon source gave a higher yield than glucose. Keywords: Volatil Fatty Acid, Fructose, Glucose, PHA, P(3HB-co-3HV), Ralstonia eutropha AbstrakBioplastik polihidroksialkanoat (PHA) adalah bioplastik dari kelompok poliester dengan sifat fisikokimia mirip dengan plastik polipropilen dari minyak bumi. Penelitian ini bertujuan untuk mempelajari pengaruh sumber karbon terhadap poduksi PHA yang dilakukan dengan proses fermentasi menggunakan Ralstonia eutropha. Ralstonia eutropha dikultivasi dalam bioreaktor batch untuk mempelajari dinamika produksi kopolimer P(3HB-co-3HV) dari sumber karbon glukosa atau fruktosa, serta mempelajari pengaruh sumber stimulator asam lemak volatil. Kondisi operasional fermentasi menggunakan bioreaktor 7 L adalah pada temperatur 30 oC dan pH 7. Konsentrasi sumber karbon glukosa atau fruktosa yang digunakan adalah 40 gr/L, dan setelah 20 jam fermentasi ditambahkan asam lemak volatil yang berfungsi sebagai stimulator dalam produksi P(3HB-co-3HV). Panen sel Ralstonia eutropha dilakukan setelah 60 jam. Hasil penelitian menunjukkan fermentasi Ralstonia eutropha dengan substrat fruktosa dan asam lemak volatil sebagai stimulator mempunyai kandungan PHA sebesar 32,78%, dengan kadar HV 11,78%. Pada pemberian substrat glukosa dan asam lemak volatil menunjukkan kandungan PHA sebesar 20,19%, dengan kadar HV 8.71%. Jadi fermentasi Ralstonia eutropha dengan menggunakan substrat fruktosa memberikan yield yang lebih tinggi dibandingkan menggunakan substrat glukosa.Kata Kunci: Asam lemak volatil, fruktosa, glukosa, PHA, P(3HB-co-3HV), Ralstonia eutropha

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Thia fatty acids with the sulfur atom in even or odd positions have opposite effects on fatty acid catabolism

Lipids ◽

10.1007/s11745-006-5085-7 ◽

2006 ◽

Vol 41 (2) ◽

pp. 169-177 ◽

Cited By ~ 9

Author(s):

Endre Dyroy ◽

Hege Wergedahl ◽

Jon Skorve ◽

Oddrun A. Gudbrandsen ◽

Jon Songstad ◽

...

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Sulfur Atom ◽

Fatty Acid Catabolism ◽

Thia Fatty Acids

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Responses of Mycobacterium tuberculosis to Growth in the Mouse Lung

Infection and Immunity ◽

10.1128/iai.73.6.3754-3757.2005 ◽

2005 ◽

Vol 73 (6) ◽

pp. 3754-3757 ◽

Cited By ~ 55

Author(s):

Eugenie Dubnau ◽

John Chan ◽

V. P. Mohan ◽

Issar Smith

Keyword(s):

Fatty Acids ◽

Mycobacterium Tuberculosis ◽

Carbon Source ◽

Real Time ◽

Real Time Pcr ◽

Mouse Lung ◽

Quantitative Real Time Pcr ◽

Promoter Trap ◽

Mouse Lungs ◽

Rna Levels

ABSTRACT Using a promoter trap, we have identified 56 Mycobacterium tuberculosis genes preferentially expressed in the mouse lung. Quantitative real-time PCR showed that RNA levels of several genes were higher from bacteria growing in mouse lungs than from broth cultures. These results support the current hypothesis that Mycobacterium tuberculosis utilizes fatty acids as a carbon source in the mouse lung.

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Differential regulation of the two-component regulatory system senX3-regX3 in Mycobacterium tuberculosis

Microbiology ◽

10.1099/mic.0.077180-0 ◽

2014 ◽

Vol 160 (6) ◽

pp. 1125-1133 ◽

Cited By ~ 11

Author(s):

Dalin Rifat ◽

Petros C. Karakousis

Keyword(s):

Mycobacterium Tuberculosis ◽

Promoter Activity ◽

Regulatory System ◽

Differential Regulation ◽

Northern Analysis ◽

Pcr Analysis ◽

Reporter System ◽

Two Component ◽

Phosphate Depletion

The highly successful pathogen Mycobacterium tuberculosis (Mtb) has evolved strategies to adapt to various stress conditions, thus promoting survival within the infected host. The two-component regulatory system (2CRS) senX3-regX3, which has been implicated in the Mtb response to inorganic phosphate depletion, is believed to behave as an auto-regulatory bicistronic operon. Unlike other 2CRS, Mtb senX3-regX3 features an intergenic region (IR) containing several mycobacterium interspersed repetitive units (MIRU) of unknown function. In this study, we used a lacZ reporter system to study the promoter activity of the 5′ untranslated region of senX3, and that of various numbers of MIRUs in the senX3-regX3 IR, during axenic Mtb growth in nutrient-rich broth, and upon exposure to growth-restricting conditions. Activity of the senX3 promoter was induced during phosphate depletion and nutrient starvation, and IR promoter activity under these conditions was directly proportional to the number of MIRUs present. Quantitative reverse transcriptase (qRT)-PCR analysis of exponentially growing Mtb revealed monocistronic transcription of senX3 and regX3, and, to a lesser degree, bicistronic transcription of the operon. In addition, we observed primarily monocistronic upregulation of regX3 during phosphate depletion of Mtb, which was confirmed by Northern analysis in wild-type Mtb and by RT-PCR in a senX3-disrupted mutant, while upregulation of regX3 in nutrient-starved Mtb was chiefly bicistronic. Our findings of differential regulation of senX3-regX3 highlight the potential regulatory role of MIRUs in the Mtb genome and provide insight into the regulatory mechanisms underlying Mtb adaptation to physiologically relevant conditions.

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