Combinatorial chemistry in nematodes: modular assembly of primary metabolism-derived building blocks

Abstract

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.

Fig. 1

Initial identification of ascarosides in nematodes. (a) C. elegans life cycle. (b) Ascarosides with potent dauer inducing activity. (c) Lipid-like ascaroside from Ascaris spp.

Fig. 2

Modular ascarosides regulating development and mediating behavioural phenotypes in C. elegans, including building blocks derived from tryptophan (red) and folate (blue) metabolism.

Fig. 3

(a) mbas#3 and hbas#3, two modular ascarosides including tigloyl and p-hydroxybenzoyl modules putatively derived from amino acid or short-chain fatty acid metabolism (red) that were identified via targeted metabolomics in mixed stage C. elegans cultures. (b) ascr#10, a male-produced hermaphrodite attractant, and osas#9, a compound produced abundantly by starved L1 larvae, incorporating succinyl (magenta) and octopamine (green) moieties.

Fig. 4

Attachment of a tryptophan-derived indole carboxy moiety to the deterrent ascr#3 results in the hermaphrodite attractant icas#3. In addition, ascr#3 is part of the male-attracting pheromone (see text).

Fig. 5

Structures of the side chain-hydroxylated dhas#18, a pheromone identified from the sour paste nematode P. redivivus, and easc#18, a component of the dauer pheromone of the entomopathogenic nematode H. bacteriophora.

Fig. 6

Ascaroside and paratoside-derived metabolites in P. pacificus. Major components of the P. pacificus exo-metabolome derived from assembly of building blocks from carbohydrate, lipid, amino acid (red), and nucleoside (blue) metabolism, as well as TCA cycle-derived succinate (magenta). Also shown is the highly conserved tRNA nucleoside, N⁶-threonylcarbamoyladenosine (t⁶A), a putative precursor of paratosides npar#1–3.

Fig. 7

The dimeric ascaroside dasc#1 promotes development of the wide (eurystomatous) mouth form that enables a predatory lifestyle consuming other nematodes.

Fig. 8

Side chain shortening of long-chain ascarosides via iterative four-step peroxisomal β-oxidation in C. elegans.^,

Fig. 9

Interaction of endocannabinoid biosynthesis with peroxisomal β-oxidation. daf-22 mutation abolishes processing of long-chain ascaroside CoA esters, whose conversion into ascaroside ethanolamides is associated with reduced EPEA and anandamide production, likely due to depletion of phosphatidylethanolamine pools. Shown in red are shunt metabolites identified in daf-22 mutant endo-metabolomes via mvaDANS.

Fig. 10

Model for the biosynthesis of the avoidance signal osas#9 in C. elegans. tbh-1 mutant worms produce large quantities of the shunt metabolite tsas#9 due to accumulation of the octopamine precursor tyramine.

Fig. 11

glas#3, an ascaroside glucosyl ester identified in the C. elegans endo-metabolome.