RESUMEN
A non-canonical biosynthetic pathway furnishing the first natural brexane-type bishomosesquiterpene (chlororaphen, C17 H28 ) was elucidated in the γ-proteobacterium Pseudomonas chlororaphis O6. A combination of genome mining, pathway cloning, in vitro enzyme assays, and NMR spectroscopy revealed a three-step pathway initiated by C10 methylation of farnesyl pyrophosphate (FPP, C15 ) along with cyclization and ring contraction to furnish monocyclic γ-presodorifen pyrophosphate (γ-PSPP, C16 ). Subsequent C-methylation of γ-PSPP by a second C-methyltransferase furnishes the monocyclic α-prechlororaphen pyrophosphate (α-PCPP, C17 ), serving as the substrate for the terpene synthase. The same biosynthetic pathway was characterized in the ß-proteobacterium Variovorax boronicumulans PHE5-4, demonstrating that non-canonical homosesquiterpene biosynthesis is more widespread in the bacterial domain than previously anticipated.
Asunto(s)
Comamonadaceae , Pseudomonas chlororaphis , Metilación , Difosfatos , Comamonadaceae/genéticaRESUMEN
Nematodes represent the most abundant group of metazoans on earth. They utilize diverse chemicals to interact with con-specific and hetero-specific organisms, and are also impacted by compounds produced by other interacting organisms. In the first part of this review we discuss how nematode-derived glycolipids modulate their behavior and development, as well as the interactions with other organisms. Furthermore, we provide a short overview about other secondary metabolites produced by nematodes that affect different life traits of free-living nematodes. In the second part of this review we discuss how different bacteria-, nematode-, and plant-derived chemicals such as volatile organic compounds, root exudates, and plant defenses regulate the interaction between entomopathogenic nematodes, their symbiotic bacteria, insect prey, predators, and plants.
RESUMEN
Comparative ascaroside profiling of Caenorhabditis nematodes using HPLC-ESI-(-)-MS/MS precursor ion scanning revealed a class of highly species-specific ascaroside dimers. Their 2- and 4-isomeric, homo- and heterodimeric structures were identified using a combination of HPLC-ESI-(+)-HR-MS/MS spectrometry and high-resolution dqf-COSY NMR spectroscopy. Structure assignments were confirmed by total synthesis of representative examples. Functional characterization using holding assays indicated that males of Caenorhabditis remanei and Caenorhabditis nigoni are exclusively retained by their conspecific ascaroside dimers, demonstrating that dimerization of conserved monomeric building blocks represents a yet undescribed mechanism that generates species-specific signaling molecules in the Caenorhabditis genus.
Asunto(s)
Caenorhabditis elegans/metabolismo , Glucolípidos/metabolismo , Animales , Cromatografía Líquida de Alta Presión/métodos , Dimerización , Espectroscopía de Resonancia Magnética/métodos , Transducción de Señal , Espectrometría de Masa por Ionización de Electrospray/métodosRESUMEN
Nematode chemosensation is a vital component of their host-seeking behavior. The globally important phytonematode Meloidogyne incognita perceives and responds (via sensory organs such as amphids and phasmids) differentially to various chemical cues emanating from the rhizosphere during the course of host finding. However, compared with the free-living worm Caenorhabditis elegans, the molecular intricacies behind the plant nematode chemotaxis are a yet-unexploited territory. In the present study, four putative chemosensory genes of M. incognita, namely, Mi-odr-1, Mi-odr-3, Mi-tax-2, and Mi-tax-4 were molecularly characterized. Mi-odr-1 mRNA was found to be expressed in the cell bodies of amphidial neurons and phasmids of M. incognita. Mi-odr-1, Mi-odr-3, Mi-tax-2, and Mi-tax-4 transcripts were highly expressed in early life stages of M. incognita, consistent with a role of these genes in host recognition. Functional characterization of Mi-odr-1, Mi-odr-3, Mi-tax-2, and Mi-tax-4 via RNA interference revealed behavioral defects in M. incognita and perturbed attraction to host roots in Pluronic gel medium. Knockdown of Mi-odr-1, Mi-odr-3, Mi-tax-2, and Mi-tax-4 resulted in defective chemotaxis of M. incognita to various volatile compounds (alcohol, ketone, aromatic compound, ester, thiazole, pyrazine), nonvolatiles of plant origin (carbohydrate, phytohormone, organic acid, amino acid, phenolic), and host root exudates in an agar-Pluronic gel-based assay plate. In addition, ascaroside-mediated signaling was impeded by downregulation of chemosensory genes. This new information that behavioral response in M. incognita is modulated by specific olfactory genes can be extended to understand chemotaxis in other nematodes.
Asunto(s)
Quimiotaxis , Tylenchoidea , Animales , Conducta Animal/fisiología , Caenorhabditis elegans/genética , Quimiotaxis/genética , Interferencia de ARN , Tylenchoidea/genética , Tylenchoidea/metabolismoRESUMEN
Chemical communication in nematodes such as the model organism Caenorhabditis elegans is modulated by a variety of glycosides based on the dideoxysugar l-ascarylose. Comparative ascaroside profiling of nematode exometabolome extracts using a GC-EIMS screen reveals that several basic components including ascr#1 (asc-C7), ascr#2 (asc-C6-MK), ascr#3 (asc-ΔC9), ascr#5 (asc-ωC3), and ascr#10 (asc-C9) are highly conserved among the Caenorhabditis. Three novel side chain hydroxylated ascaroside derivatives were exclusively detected in the distantly related C. nigoni and C. afra. Molecular structures of these species-specific putative signaling molecules were elucidated by NMR spectroscopy and confirmed by total synthesis and chemical correlations. Biological activities were evaluated using attraction assays. The identification of (ω)- and (ω - 2)-hydroxyacyl ascarosides demonstrates how GC-EIMS-based ascaroside profiling facilitates the detection of novel ascaroside components and exemplifies how species-specific hydroxylation of ascaroside aglycones downstream of peroxisomal ß-oxidation increases the structural diversity of this highly conserved class of nematode signaling molecules.
Asunto(s)
Caenorhabditis elegans/metabolismo , Cromatografía de Gases y Espectrometría de Masas/métodos , Peroxisomas/metabolismo , Espectrometría de Masa por Ionización de Electrospray/métodos , Animales , Hidroxilación , Espectroscopía de Resonancia Magnética/métodos , Oxidación-ReducciónRESUMEN
NMR chemical profiling of a Laurenciella sp. using a computerized method developed in our laboratory resulted in the identification of five new compounds (1-5) and 17 known compounds, among which 3-(E)-laurenyne represented by far the most abundant metabolite. Compounds 1 to 5 were isolated and fully characterized by detailed spectroscopic analysis. The absolute configuration and structural features of compound 1 were determined by single-crystal X-ray diffraction analysis. Compounds 1 to 4 are 12-membered cyclic ether acetogenins that are present in solution as interconverting conformers exhibiting an (aR) configuration of the bromoallene unit together with an S configuration at C-4. Among these, compound 3 is the first obtusallene derivative with bromine substituents at both the C-7 and C-12 positions. Compound 5 is an acetogenin bearing a [5.5.1]bicyclotridecane ring system. A plausible biosynthetic route to 1-4 is proposed.
RESUMEN
Chemical communication in nematodes has been known for over half a century, but the underlying molecular basis remained largely elusive. Recent advances in analytical techniques facilitated the characterization of a modular glycolipid library based on the dideoxysugar L-ascarylose, which modulates behavior and development in the model organism C. elegans. Ascaroside signaling is highly conserved in nematodes and represents a key factor in nematode chemical ecology. Ascaroside biosynthesis depends on the co-option of the peroxisomal ß-oxidation cycle and in addition integrates a large diversity of additional building blocks derived from various primary metabolic pathways to give rise to species-specific modular assemblies, thus, transcending the concept of strictly segregated primary versus secondary metabolism.
Asunto(s)
Comunicación Animal , Caenorhabditis elegans/fisiología , Glucolípidos/fisiología , Animales , Comunicación , HexosasRESUMEN
Nematodes such as the model organism Caenorhabditis elegans produce various homologous series of l-ascarylose-derived glycolipids called ascarosides, which include several highly potent signals in intra and interspecies communication as well as cross-kingdom interactions. Given their low concentrations and large number of structurally similar components, mass spectrometric screens based on high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) are commonly employed for ascaroside detection and quantification. Here, we describe a complementary gas chromatography-electron ionization mass spectrometry (GC-EIMS) screen that utilizes an ascarylose-derived K1-fragment ion signal at m/z 130.1 [C6H14OSi]+â to highlight known as well as yet unidentified ascaroside components in TMS-derivatized crude nematode exometabolome extracts. GC-EIMS-based ascaroside profiling of wild-type and mutant C. elegans facilitates the analysis of all basic ascarosides using the same ionization technique while providing excellent resolution for the complete homologous series with side chains ranging from 3 to 33 carbons. Combined screening for m/z 130.1 along with side chain-specific J1 [M - 173]+ and J2 [M - 291]+ fragment ions, as well as additional characteristic marker ions from α-cleavage, enables convenient structure assignment of ca. 200 components from wild-type and peroxisomal ß-oxidation mutants including (ω - 1)-linked acyl, enoyl, ß-hydroxyacyl, and 2-ketoalkyl ascarosides along with their (ω)-linked or α-methyl isomers and ethanolamide derivatives, as well as 2-hydroxyalkyl ascarosides. Given the widespread availability of GC-MS and its increasing popularity in metabolomics, this method will promote the identification of ascarosides in C. elegans and other nematodes.
Asunto(s)
Caenorhabditis elegans/metabolismo , Cromatografía de Gases y Espectrometría de Masas/métodos , Glucolípidos/análisis , Metaboloma , Animales , Glucolípidos/metabolismo , MetabolómicaRESUMEN
UNLABELLED: The tetrachloroethene (PCE)-respiring bacterium Sulfurospirillum multivorans produces a unique cobamide, namely, norpseudo-B12, which, in comparison to other cobamides, e.g., cobalamin and pseudo-B12, lacks the methyl group in the linker moiety of the nucleotide loop. In this study, the protein SMUL_1544 was shown to be responsible for the formation of the unusual linker moiety, which is most probably derived from ethanolamine-phosphate (EA-P) as the precursor. The product of the SMUL_1544 gene successfully complemented a Salmonella enterica ΔcobD mutant. The cobD gene encodes an l-threonine-O-3-phosphate (l-Thr-P) decarboxylase responsible for the synthesis of (R)-1-aminopropan-2-ol O-2-phosphate (AP-P), required specifically for cobamide biosynthesis. When SMUL_1544 was produced in the heterologous host lacking CobD, norpseudo-B12 was formed, which pointed toward the formation of EA-P rather than AP-P. Guided cobamide biosynthesis experiments with minimal medium supplemented with l-Thr-P supported cobamide biosynthesis in S. enterica producing SMUL_1544 or S. multivorans Under these conditions, both microorganisms synthesized pseudo-B12 This observation indicated a flexibility in the SMUL_1544 substrate spectrum. From the formation of catalytically active PCE reductive dehalogenase (PceA) in S. multivorans cells producing pseudo-B12, a compatibility of the respiratory enzyme with the cofactor was deduced. This result might indicate a structural flexibility of PceA in cobamide binding. Feeding of l-[3-(13)C]serine to cultures of S. multivorans resulted in isotope labeling of the norpseudo-B12 linker moiety, which strongly supports the hypothesis of EA-P formation from l-serine-O-phosphate (l-Ser-P) in this organism. IMPORTANCE: The identification of the gene product SMUL_1544 as a putative l-Ser-P decarboxylase involved in norcobamide biosynthesis in S. multivorans adds a novel module to the assembly line of cobamides (complete corrinoids) in prokaryotes. Selected cobamide-containing enzymes (e.g., reductive dehalogenases) showed specificity for their cobamide cofactors. It has recently been proposed that the structure of the linker moiety of norpseudo-B12 and the mode of binding of the EA-P linker to the PceA enzyme reflect the high specificity of the enzyme for its cofactor. Data reported herein do not support this idea. In fact, norpseudo-B12 was functional in the cobamide-dependent methionine biosynthesis of S. enterica, raising questions about the role of norcobamides in nature.
Asunto(s)
Proteínas Bacterianas/metabolismo , Cobamidas/biosíntesis , Epsilonproteobacteria/metabolismo , Regulación Bacteriana de la Expresión Génica/fisiología , Tetracloroetileno/metabolismo , Proteínas Bacterianas/genética , Cobamidas/química , Cobamidas/metabolismo , Estructura MolecularRESUMEN
The indole ascarosides (icas) represent a highly potent class of nematode-derived modular signalling components that integrate structural inputs from amino acid, carbohydrate, and fatty acid metabolism. Comparative analysis of the crude exo-metabolome of hermaphroditic Caenorhabditis briggsae using a highly sensitive mass spectrometric screen reveals an indole ascaroside blend dominated by two new components. The structures of isolated icas#2 and icas#6.2 were determined by NMR spectroscopy and confirmed by total synthesis and chemical correlation. Low atto- to femtomolar amounts of icas#2 and icas#6.2 act in synergism to attract males indicating a function as sex pheromone. Comparative analysis of 14 Caenorhabditis species further demonstrates that species-specific indole ascaroside biosynthesis is highly conserved in the Elegans group. Functional characterization of the dominating indole ascarosides icas#2, icas#3, and icas#9 reveals a high degree of species-specificity and considerable variability with respect to gender-specificity, thus, confirming that indole ascarosides modulate different biological functions within the Elegans group. Although the nematode response was usually most pronounced towards conspecific signals, Caenorhabditis brenneri, the only species of the Elegans group that does not produce any indole ascarosides, exhibits a robust response to icas#2 suggesting the potential for interspecies interactions.
Asunto(s)
Caenorhabditis/química , Glicósidos/metabolismo , Indoles/metabolismo , Atractivos Sexuales/análisis , Transducción de Señal , Animales , Glicósidos/química , Indoles/química , Espectrometría de Masas , Conformación Molecular , Especificidad de la EspecieRESUMEN
Juveniles of the leaf beetles in subtribe Chrysomelina have efficient defense strategies against predators. When disturbed, they transiently expose volatile deterrents in large droplets from nine pairs of defensive glands on their back. Here, we report on an additional line of defense consisting of the non-volatile isoxazolin-5-one glucoside and its 3-nitropropanoyl ester in the larval hemolymph. Because isoxazolin-5-one derivatives were not detectable in related leaf beetle taxa, they serve as a diagnostic marker for the Chrysomelina subtribe. Conjugation of isotopically labelled 3-nitropropionic acid to isoxazolin-5-one glucoside in vivo demonstrates its function as a carrier for the 3-nitropropanoyl esters. The previous identification of characteristic glucosides as precursors of the volatile deterrents underlines the general importance of glucosides for sequestration from food plants, and the subsequent transport in the hemolymph to the defense system. The combination of repellent volatiles with non-volatile toxic compounds in the hemolymph has the potential to create synergistic effects since the odorant stimulus may help predators learn to avoid some foods. The combination of the two defense lines has the advantage, that the hemolymph toxins provide reliable and durable protection, while the repellents may vary after a host plant change.
Asunto(s)
Escarabajos/fisiología , Hemolinfa/metabolismo , Nitrocompuestos/metabolismo , Propionatos/metabolismo , Animales , Escarabajos/metabolismo , Ésteres , Filogenia , Conducta PredatoriaRESUMEN
The nematode Caenorhabditis elegans was the first animal to have its genome fully sequenced and has become an important model organism for biomedical research. However, like many other animal model systems, its metabolome remained largely uncharacterized, until recent investigations demonstrated the importance of small molecule-based signalling cascades for virtually every aspect of nematode biology. These studies have revealed that nematodes are amazingly skilled chemists: using simple building blocks from conserved primary metabolism and a strategy of modular assembly, C. elegans and other nematode species create complex molecular architectures to regulate their development and behaviour. These nematode-derived modular metabolites (NDMMs) are based on the dideoxysugars ascarylose or paratose, which serve as scaffolds for attachment of moieties from lipid, amino acid, carbohydrate, citrate, and nucleoside metabolism. Mutant screens and comparative metabolomics based on NMR spectroscopy and MS have so-far revealed several 100 different ascarylose ("ascarosides") and a few paratose ("paratosides") derivatives, many of which represent potent signalling molecules that can be active at femtomolar levels, regulating development, behaviour, body shape, and many other life history traits. NDMM biosynthesis appears to be carefully regulated as assembly of different modules proceeds with very high specificity. Preliminary biosynthetic studies have confirmed the primary metabolism origin of some NDMM building blocks, whereas the mechanisms that underlie their highly specific assembly are not understood. Considering their functions and biosynthetic origin, NDMMs represent a new class of natural products that cannot easily be classified as "primary" or "secondary". We believe that the identification of new variants of primary metabolism-derived structures that serve important signalling functions in C. elegans and other nematodes provides a strong incentive for a comprehensive re-analysis of metabolism in higher animals, including humans.
Asunto(s)
Caenorhabditis elegans/metabolismo , Nematodos/metabolismo , Animales , Caenorhabditis elegans/química , Espectroscopía de Resonancia Magnética , Metabolómica , Estructura Molecular , Nematodos/química , Transducción de Señal , Relación Estructura-ActividadRESUMEN
The nematode C. elegans is an important model for the study of social behaviors. Recent investigations have shown that a family of small molecule signals, the ascarosides, controls population density sensing and mating behavior. However, despite extensive studies of C. elegans aggregation behaviors, no intraspecific signals promoting attraction or aggregation of wild-type hermaphrodites have been identified. Using comparative metabolomics, we show that the known ascarosides are accompanied by a series of derivatives featuring a tryptophan-derived indole moiety. Behavioral assays demonstrate that these indole ascarosides serve as potent intraspecific attraction and aggregation signals for hermaphrodites, in contrast to ascarosides lacking the indole group, which are repulsive. Hermaphrodite attraction to indole ascarosides depends on the ASK amphid sensory neurons. Downstream of the ASK sensory neuron, the interneuron AIA is required for mediating attraction to indole ascarosides instead of the RMG interneurons, which previous studies have shown to integrate attraction and aggregation signals from ASK and other sensory neurons. The role of the RMG interneuron in mediating aggregation and attraction is thought to depend on the neuropeptide Y-like receptor NPR-1, because solitary and social C. elegans strains are distinguished by different npr-1 variants. We show that indole ascarosides promote attraction and aggregation in both solitary and social C. elegans strains. The identification of indole ascarosides as aggregation signals reveals unexpected complexity of social signaling in C. elegans, which appears to be based on a modular library of ascarosides integrating building blocks derived from lipid ß-oxidation and amino-acid metabolism. Variation of modules results in strongly altered signaling content, as addition of a tryptophan-derived indole unit to repellent ascarosides produces strongly attractive indole ascarosides. Our findings show that the library of ascarosides represents a highly developed chemical language integrating different neurophysiological pathways to mediate social communication in C. elegans.
Asunto(s)
Conducta Animal/efectos de los fármacos , Caenorhabditis elegans/fisiología , Bibliotecas de Moléculas Pequeñas/farmacología , Conducta Social , Animales , Caenorhabditis elegans/efectos de los fármacos , Caenorhabditis elegans/metabolismo , Glucolípidos/química , Glucolípidos/farmacología , Glicósidos/análisis , Glicósidos/metabolismo , Glicósidos/farmacología , Organismos Hermafroditas/efectos de los fármacos , Organismos Hermafroditas/metabolismo , Metaboloma/efectos de los fármacos , Metaboloma/fisiología , Metabolómica/métodos , Modelos Biológicos , Feromonas/química , Feromonas/metabolismo , Feromonas/farmacología , Transducción de Señal/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/metabolismo , Triptófano/química , Triptófano/metabolismo , Triptófano/farmacologíaRESUMEN
A novel procedure for the synthesis of isoxazolin-5-one glycosides starting from unprotected carbohydrates is described. The substrate scope of the one-pot synthetic protocol was explored using D-configured glucose, xylose, maltose, fructose, ribose and 2-deoxyribose. Naturally occurring 2-(ß-D-glucopyranosyl)-3-isoxazolin-5-one and four novel isoxazolin-5-one glycosides derived from xylose, maltose and fructose were synthesized and purified by flash chromatography. The compounds were characterized in terms of chemical structure, photophysical properties as well as pH stability. The photohydrolysis rates of the synthesized glycosides were compared with uridine as a standard to determine the quantum yields for the photoreactions in water.
Asunto(s)
Glicósidos/química , Glicósidos/síntesis química , Isoxazoles/química , Luz , Técnicas de Química Sintética , Estabilidad de Medicamentos , Concentración de Iones de HidrógenoRESUMEN
Nematodes use an extensive chemical language based on glycosides of the dideoxysugar ascarylose for developmental regulation (dauer formation), male sex attraction, aggregation, and dispersal. However, no examples of a female- or hermaphrodite-specific sex attractant have been identified to date. In this study, we investigated the pheromone system of the gonochoristic sour paste nematode Panagrellus redivivus, which produces sex-specific attractants of the opposite sex. Activity-guided fractionation of the P. redivivus exometabolome revealed that males are strongly attracted to ascr#1 (also known as daumone), an ascaroside previously identified from Caenorhabditis elegans hermaphrodites. Female P. redivivus are repelled by high concentrations of ascr#1 but are specifically attracted to a previously unknown ascaroside that we named dhas#18, a dihydroxy derivative of the known ascr#18 and an ascaroside that features extensive functionalization of the lipid-derived side chain. Targeted profiling of the P. redivivus exometabolome revealed several additional ascarosides that did not induce strong chemotaxis. We show that P. redivivus females, but not males, produce the male-attracting ascr#1, whereas males, but not females, produce the female-attracting dhas#18. These results show that ascaroside biosynthesis in P. redivivus is highly sex-specific. Furthermore, the extensive side chain functionalization in dhas#18, which is reminiscent of polyketide-derived natural products, indicates unanticipated biosynthetic capabilities in nematodes.
Asunto(s)
Nematodos/metabolismo , Atractivos Sexuales/metabolismo , Animales , Quimiotaxis/efectos de los fármacos , Femenino , Espectroscopía de Resonancia Magnética/métodos , Masculino , Metabolómica , Modelos Biológicos , Modelos Químicos , Oxígeno/química , Peroxisomas/metabolismo , Feromonas/metabolismo , Conducta Sexual Animal/efectos de los fármacos , Conducta Sexual Animal/fisiología , Transducción de SeñalRESUMEN
Green leaf volatiles (GLVs) consisting of six-carbon aldehydes, alcohols, and their esters, are biosynthesized through the action of fatty acid hydroperoxide lyase (HPL), which uses fatty acid hydroperoxides as substrates. GLVs form immediately after disruption of plant leaf tissues by herbivore attacks and mechanical wounding and play a role in defense against attackers that attempt to invade through the wounds. The fates and the physiological significance of the counterparts of the HPL reaction, the 12/10-carbon oxoacids that are formed from 18/16-carbon fatty acid 13-/11-hydroperoxides, respectively, are largely unknown. In this study, we detected monogalactosyl diacylglycerols (MGDGs) containing the 12/10-carbon HPL products in disrupted leaf tissues of Arabidopsis, cabbage, tobacco, tomato, and common bean. They were identified as an MGDG containing 12-oxo-9-hydroxy-(E)-10-dodecenoic acid and 10-oxo-7-hydroxy-(E)-8-decenoic acid and an MGDG containing two 12-oxo-9-hydroxy-(E)-10-dodecenoic acids as their acyl groups. Analyses of Arabidopsis mutants lacking HPL indicated that these MGDGs were formed enzymatically through an active HPL reaction. Thus, our results suggested that in disrupted leaf tissues, MGDG-hydroperoxides were cleaved by HPL to form volatile six-carbon aldehydes and non-volatile 12/10-carbon aldehyde-containing galactolipids. Based on these results, we propose a novel oxylipin pathway that does not require the lipase reaction to form GLVs.
Asunto(s)
Arabidopsis/metabolismo , Ácidos Grasos Monoinsaturados/metabolismo , Galactolípidos/metabolismo , Aceites Volátiles/metabolismo , Oxilipinas/metabolismo , Hojas de la Planta/metabolismo , Aldehído-Liasas/genética , Aldehído-Liasas/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Brassica/genética , Brassica/metabolismo , Sistema Enzimático del Citocromo P-450/genética , Sistema Enzimático del Citocromo P-450/metabolismo , Galactolípidos/genética , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismo , Phaseolus/genética , Phaseolus/metabolismo , Hojas de la Planta/genética , Nicotiana/genética , Nicotiana/metabolismoRESUMEN
The ascarosides, small-molecule signals derived from combinatorial assembly of primary metabolism-derived building blocks, play a central role in Caenorhabditis elegans biology and regulate many aspects of development and behavior in this model organism as well as in other nematodes. Using HPLC-MS/MS-based targeted metabolomics, we identified novel ascarosides incorporating a side chain derived from succinylation of the neurotransmitter octopamine. These compounds, named osas#2, osas#9, and osas#10, are produced predominantly by L1 larvae, where they serve as part of a dispersal signal, whereas these ascarosides are largely absent from the metabolomes of other life stages. Investigating the biogenesis of these octopamine-derived ascarosides, we found that succinylation represents a previously unrecognized pathway of biogenic amine metabolism. At physiological concentrations, the neurotransmitters serotonin, dopamine, and octopamine are converted to a large extent into the corresponding succinates, in addition to the previously described acetates. Chemically, bimodal deactivation of biogenic amines via acetylation and succinylation parallels posttranslational modification of proteins via acetylation and succinylation of L-lysine. Our results reveal a small-molecule connection between neurotransmitter signaling and interorganismal regulation of behavior and suggest that ascaroside biosynthesis is based in part on co-option of degradative biochemical pathways.
Asunto(s)
Aminas Biogénicas/metabolismo , Caenorhabditis elegans/metabolismo , Octopamina/química , Agonistas alfa-Adrenérgicos/química , Animales , Conducta Animal , Cromatografía Líquida de Alta Presión , Dopamina/metabolismo , Glicósidos/química , Espectrometría de Masas , Neurotransmisores/metabolismo , Feromonas/metabolismo , Serotonina/metabolismo , Transducción de Señal , Succinatos/químicaRESUMEN
A novel synthetic route was developed for the construction of isoxazolin-5-one glucosides using a cascade reaction. An X-ray crystal structure analysis of a isoxazolin-5-one glucoside confirmed the structure and stereochemistry of the heterocycle. The properties of the α- and ß-anomers of the isoxazolin-5-one glucosides were compared. The first synthesis of 2-[6'-(3â³-nitropropanoyl)-ß-D-glucopyranosyl]-3-isoxazolin-5-one was realized by direct enzymatic esterification without need of further protective groups.
Asunto(s)
Glucósidos/síntesis química , Isoxazoles/síntesis química , Cristalografía por Rayos X , Glucósidos/química , Isoxazoles/química , Modelos Moleculares , Conformación MolecularRESUMEN
In the model organism Caenorhabditis elegans, a family of endogenous small molecules, the ascarosides function as key regulators of developmental timing and behavior that act upstream of conserved signaling pathways. The ascarosides are based on the dideoxysugar ascarylose, which is linked to fatty-acid-like side chains of varying lengths derived from peroxisomal ß-oxidation. Despite the importance of ascarosides for many aspects of C. elegans biology, knowledge of their structures, biosynthesis, and homeostasis remains incomplete. We used an MS/MS-based screen to profile ascarosides in C. elegans wild-type and mutant metabolomes, which revealed a much greater structural diversity of ascaroside derivatives than previously reported. Comparison of the metabolomes from wild-type and a series of peroxisomal ß-oxidation mutants showed that the enoyl CoA-hydratase MAOC-1 serves an important role in ascaroside biosynthesis and clarified the functions of two other enzymes, ACOX-1 and DHS-28. We show that, following peroxisomal ß-oxidation, the ascarosides are selectively derivatized with moieties of varied biogenetic origin and that such modifications can dramatically affect biological activity, producing signaling molecules active at low femtomolar concentrations. Based on these results, the ascarosides appear as a modular library of small-molecule signals, integrating building blocks from three major metabolic pathways: carbohydrate metabolism, peroxisomal ß-oxidation of fatty acids, and amino acid catabolism. Our screen further demonstrates that ascaroside biosynthesis is directly affected by nutritional status and that excretion of the final products is highly selective.