The laboratory of bioanalytical chemistry develops novel analytical techniques to decipher the ecology and evolution of small molecule signaling and secondary metabolism in nematodes.

Nematode Chemical Ecology and Secondary Metabolism

Nematodes (roundworms) represent the largest group of animals on earth, both in total number of individuals and the presumed number of (largely undescribed) species. Nematodes occupy almost every biological niche in terrestrial and aquatic habitats and utilize an extensive diversity of food sources, ranging from free-living bacterivorous and fungivorous species, to entomopathogenic species and plant or animal parasites. The free-living bacterivorous Caenorhabditis elegans (Rhabditidae) represents an extremely successful model organism in molecular and developmental biology, but its secondary metabolism and chemical ecology is poorly understood. Chemical signaling in nematodes has been known since the 1960s but molecular structures have remained elusive for several decades.

During the last years, research with C. elegans and a few other species along with advances in analytical techniques has revealed that chemical signaling in nematodes involves a homologous library of glycolipids called ascarosides, that is based on the 3,6-dideoxy-arabino-hexopyranose sugar L-ascarylose and fatty acid-derived aglycones. Ascaroside signaling is highly conserved in nematodes and modulates developmental plasticity and a variety of sex-specific and social behaviors in intra- and interspecies as well as cross kingdom interactions, indicating that ascarosides represents key regulators in nematode chemical ecology. In addition to ascaroside-type glycolipids, nematodes also produce an unexpected diversity of species-specific and surprisingly complex modular metabolites, which combine structural units from various primary biosynthetic pathways and whose biological functions are largely unknown. Biosynthesis of secondary metabolites via a modular assembly of ubiquitous building blocks derived from primary metabolism requires only a minimum number of additional genes (in contrast to a highly dedicated complex biosynthetic gene clusters) while potentially creating a vast chemical space. Furthermore, considering that bacterivorous nematodes naturally co-occur with a large diversity of microorganisms, many of which are known to produce several highly potent secondary metabolites, the assembly of nematode derived modular metabolites might also serve as a detoxification pathway in nematode bacteria interactions.   

At the laboratory of bioanalytical chemistry, our multidisciplinary research combines state of the art chemical analysis, total synthesis, biosynthetic studies, and functional characterization of nematode-derived secondary metabolites in order to decipher the molecular basis of nematode chemical ecology and secondary metabolism.

Chemical Analysis

Nematode-derived signaling molecules exhibit biological activities in extremely small amounts, down to the attomol (10^–18 mol) range, and are released into a complex aqueous matrix of highly dominating primary and secondary metabolites, which renders their detection and identification quite challenging. Consequently, we aim to develop novel mass spectrometric screens that facilitate the selective detection of known as well as yet unidentified ascaroside components and employ these techniques to characterize nematode metabolomes. Following a phylometabolomic approach, we utilize comparative metabolomics and identify novel molecular structures of both common and highly species-specific putative signaling components, to decipher the ecology and evolution of ascaroside signaling in Caenorhabditis and other nematode genera. Target compounds are subsequently enriched from metabolome extracts using chromatographic techniques and their structures identified by a combination of mass spectrometric and NMR spectroscopic techniques.

Total Synthesis

Due to the limitations with respect to total amounts and absolute purities of small molecule signals that can be isolated from nematode metabolome extracts, their structure assignments are often based on limited analytical datasets consisting of one and two-dimensional ¹H NMR spectra and HR-MS/MS data of only the most abundant components. Consequently, in order to confirm our structure assignments the total synthesis of selected target compounds represents a fundamentally important part of our research. Pure compounds generated by total synthesis also serve as authentic reference standards for their quantification in nematode metabolome extracts and the development of novel analytical screens to identify homologs and other structurally related derivatives. Finally, the synthetic materials are employed for the characterization of their biological functions in biologically relevant concentrations.

Biosynthesis

In addition to our research in analytical and synthetic chemistry, we also strive to elucidate metabolic pathways involved in the biosynthesis of nematode-derived secondary metabolites by incorporation of stable isotope-labeled putative precursors: Furthermore, we study secondary metabolism in nematodes by comparative analysis of wild type and mutant metabolomes, which also facilitates the functional characterization of the corresponding genes. To unravel the biological significance of individual metabolites and putative signaling molecules we analyze how their production depends on nematode gender and development and how it is influenced by environmental factors such as temperature, population density, food source, and food availability, which puts individual components into an ecological context.

Functional Characterization

To characterize the biological functions of putative small molecule signals we employ a diverse array of bioassays to study nematode development and behavior. In this context, we are particularly interested in intra- and interspecies interactions as well as cross kingdom interactions with microorganisms, host organisms, and nematode predators.