The Biosynthesis of Ascarosides in Caenorhabditis elegans

Ascarosides comprise a family of small signaling molecules that have been shown to regulate important events and behaviors in the life history of the nematode Caenorhabditis elegans. Although the different roles of individual ascarosides appear to be determined by the variances in chemical structure, the mechanisms by which ascarosides are synthesized as well as the locations in which ascarosides are produced within the worm are largely unknown. In this thesis, we examined ascaroside production in the intestine, hypodermis, and body wall muscle of the worm by driving the expression of the protein DAF-22 under different tissue-specific gene promoters. While the body wall muscle and hypodermis are capable of synthesizing ascarosides, the intestine appears to be the major site of pheromone production. Additionally, we found through transgenic rescue and HPLC-MS analysis, that the acyl-CoA synthetase ACS-7 plays a significant role in the addition of moieties derived from primary metabolic pathways to the 4’-position of the ascarylose sugar core of ascr#9.</p

Metabolomic “Dark Matter” Dependent on Peroxisomal β-Oxidation in Caenorhabditis elegans

Peroxisomal β-oxidation (pβo) is a highly conserved fat metabolism pathway involved in the biosynthesis of diverse signaling molecules in animals and plants. In Caenorhabditis elegans, pβo is required for the biosynthesis of the ascarosides, signaling molecules that control development, lifespan, and behavior in this model organism. Via comparative mass spectrometric analysis of pβo mutants and wildtype, we show that pβo in C. elegans and the satellite model P. pacificus contributes to life stage-specific biosynthesis of several hundred previously unknown metabolites. The pβo-dependent portion of the metabolome is unexpectedly diverse, e.g., intersecting with nucleoside and neurotransmitter metabolism. Cell type-specific restoration of pβo in pβo-defective mutants further revealed that pβo-dependent submetabolomes differ between tissues. These results suggest that interactions of fat, nucleoside, and other primary metabolism pathways can generate structural diversity reminiscent of that arising from combinatorial strategies in microbial natural product biosynthesis

Biosynthesis of Modular Ascarosides in C. elegans

The nematode Caenorhabditis elegans uses simple building blocks from primary metabolism and a strategy of modular assembly to build a great diversity of signaling molecules, the ascarosides, which function as a chemical language in this model organism. In the ascarosides, the dideoxysugar ascarylose serves as a scaffold to which diverse moieties from lipid, amino acid, neurotransmitter, and nucleoside metabolism are attached. However, the mechanisms that underlie the highly specific assembly of ascarosides are not understood. We show that the acyl-CoA synthetase ACS-7, which localizes to lysosome-related organelles, is specifically required for the attachment of different building blocks to the 4′-position of ascr#9. We further show that mutants lacking lysosome-related organelles are defective in the production of all 4′-modified ascarosides, thus identifying the waste disposal system of the cell as a hotspot for ascaroside biosynthesis

Metabolomic “Dark Matter” Dependent on Peroxisomal β-Oxidation in Caenorhabditis elegans

Peroxisomal β-oxidation (pβo) is a highly conserved fat metabolism pathway involved in the biosynthesis of diverse signaling molecules in animals and plants. In Caenorhabditis elegans, pβo is required for the biosynthesis of the ascarosides, signaling molecules that control development, lifespan, and behavior in this model organism. Via comparative mass spectrometric analysis of pβo mutants and wildtype, we show that pβo in C. elegans and the satellite model P. pacificus contributes to life stage-specific biosynthesis of several hundred previously unknown metabolites. The pβo-dependent portion of the metabolome is unexpectedly diverse, e.g., intersecting with nucleoside and neurotransmitter metabolism. Cell type-specific restoration of pβo in pβo-defective mutants further revealed that pβo-dependent submetabolomes differ between tissues. These results suggest that interactions of fat, nucleoside, and other primary metabolism pathways can generate structural diversity reminiscent of that arising from combinatorial strategies in microbial natural product biosynthesis

Biology and genome of a newly discovered sibling species of Caenorhabditis elegans

A ‘sibling’ species of the model organism Caenorhabditis elegans has long been sought for use in comparative analyses that would enable deep evolutionary interpretations of biological phenomena. Here, we describe the first sibling species of C. elegans, C. inopinata n. sp., isolated from fig syconia in Okinawa, Japan. We investigate the morphology, developmental processes and behaviour of C. inopinata, which differ significantly from those of C. elegans. The 123-Mb C. inopinata genome was sequenced and assembled into six nuclear chromosomes, allowing delineation of Caenorhabditis genome evolution and revealing unique characteristics, such as highly expanded transposable elements that might have contributed to the genome evolution of C. inopinata. In addition, C. inopinata exhibits massive gene losses in chemoreceptor gene families, which could be correlated with its limited habitat area. We have developed genetic and molecular techniques for C. inopinata; thus C. inopinata provides an exciting new platform for comparative evolutionary studies

Tissue-specific ascaroside production in the nematode Caenorhabditis elegans

Ascarosides are a family of small mols. used as signals in a variety of important behaviors in C. elegans and other nematodes; these include mate attraction, aggression, repulsion, and entry into dauer diapause under stressful conditions. Structurally, ascarosides are modular glycosides of the dideoxysugar ascarylose. Small structural changes in the fatty add moiety attached to the first position of the sugar as well as the addn. of other metabolically-derived components to the fourth position have been shown to elicit different behaviors. DAF-22, a thiolase responsible for the last step of peroxisomal beta-oxidn. of the ascaroside lipid side chain, is necessary for the prodn. of biol. active ascarosides and is normally expressed in the intestine, body wall muscle, and hypodermis of the worm. After transgenically rescuing the expression of DAF-22 in each tissue individually in a DAF-22 knockout background, HPLC-MS anal. revealed that all three tissues are capable of producing ascarosides, but in different quantities. Furthermore, the expression of DAF-22 in any of the three tissues rescues the ability of the worm to enter dauer diapause, but to varying degrees. By studying these site-specific activity relationships, we highlight the roles of multiple tissues in worm communication via ascarosides

Metabolomic “Dark Matter” Dependent on Peroxisomal β‑Oxidation in <i>Caenorhabditis elegans</i>

Peroxisomal β-oxidation (pβo) is a highly conserved fat metabolism pathway involved in the biosynthesis of diverse signaling molecules in animals and plants. In <i>Caenorhabditis elegans</i>, pβo is required for the biosynthesis of the ascarosides, signaling molecules that control development, lifespan, and behavior in this model organism. Via comparative mass spectrometric analysis of pβo mutants and wildtype, we show that pβo in <i>C. elegans</i> and the satellite model <i>P. pacificus</i> contributes to life stage-specific biosynthesis of several hundred previously unknown metabolites. The pβo-dependent portion of the metabolome is unexpectedly diverse, e.g., intersecting with nucleoside and neurotransmitter metabolism. Cell type-specific restoration of pβo in pβo-defective mutants further revealed that pβo-dependent submetabolomes differ between tissues. These results suggest that interactions of fat, nucleoside, and other primary metabolism pathways can generate structural diversity reminiscent of that arising from combinatorial strategies in microbial natural product biosynthesis

Metabolomic “Dark Matter” Dependent on Peroxisomal β‑Oxidation in <i>Caenorhabditis elegans</i>

Peroxisomal β-oxidation (pβo) is a highly conserved fat metabolism pathway involved in the biosynthesis of diverse signaling molecules in animals and plants. In <i>Caenorhabditis elegans</i>, pβo is required for the biosynthesis of the ascarosides, signaling molecules that control development, lifespan, and behavior in this model organism. Via comparative mass spectrometric analysis of pβo mutants and wildtype, we show that pβo in <i>C. elegans</i> and the satellite model <i>P. pacificus</i> contributes to life stage-specific biosynthesis of several hundred previously unknown metabolites. The pβo-dependent portion of the metabolome is unexpectedly diverse, e.g., intersecting with nucleoside and neurotransmitter metabolism. Cell type-specific restoration of pβo in pβo-defective mutants further revealed that pβo-dependent submetabolomes differ between tissues. These results suggest that interactions of fat, nucleoside, and other primary metabolism pathways can generate structural diversity reminiscent of that arising from combinatorial strategies in microbial natural product biosynthesis