The Search for Novel Anthelmintic Targets: Characterizing Alternative Metabolic Pathways in Caenorhabditis elegans

Around a quarter of the human population is infected by parasitic helminths which also infect and place a large economic burden on the agricultural industry. Unfortunately anthelmintic resistance is a growing problem. Helminths survive long periods of low to no oxygen in their hosts, but their anaerobic metabolism has yet to be fully characterized. The goal of this project has been to characterize the anaerobic metabolism of C. elegans, a roundworm closely related to a large class of parasitic worms, by combining systematic metabolomics with a structural biology approach in order to find selective drug targets. Fumarate reduction has been previously identified as the primary source of ATP for C.elegans as well as many helminths in low oxygen environments, such as during host infection. Rhodoquinone (RQ) is an essential small molecule which is closely related to ubiquinone (UQ). While UQ is essential for the canonical, aerobic electron transport chain (ETC) which generates the majority of ATP in most animals, RQ is essential for fumarate reduction and is the fulcrum of anaerobic metabolic pathways more generally, here referred to as RQ-dependent metabolism (RQDM). In this thesis, I established a new methodology for inducing fumarate reduction in C.elegans with cyanide (KCN) which increases ease of study. I have found that wild-type C.elegans treated with KCN have the elevated succinate levels characteristic of fumarate reduction in helminths. In RQ-deficient mutants treated with KCN, succinate levels are reduced while its precursors, malate and fumarate, are elevated. This confirms the structure of the hypothesized fumarate reduction pathway in a whole animal. This finding was replicated when Complex I, which is likewise required for fumarate reduction, was inhibited by rotenone. By combining statistical analyses of metabolites in animals where fumarate reduction was disrupted in different ways with a list of known RQDM enzymes, I identified twelve (12) genes which are required for RQDM, four (4) of which have helminth-specific residues near their active site and one (1) of which is not found in hosts. These discoveries provide a new list of high-priority candidates for anthelmintic drug targets.

Department dc:contributor.department

Molecular Genetics

Year dc:date.issued

2023

Author dc:creator

• Lautens, Margot Jeanine

Advisor dc:contributor.advisor

• Fraser, Andrew

• ANOVA
• Anthelmintics
• C.elegans
• Helminths
• LC-MS
• PEPCK

Statement dc:rights

• Attribution 4.0 International

Licence dc:rights.uri

• http://creativecommons.org/licenses/by/4.0/

Handle dc:identifier.uri

http://hdl.handle.net/1807/130367

OAI identifier oai:identifier

oai:utoronto.scholaris.ca:1807/130367