Resolving the unique mechanism of endocytosis in Giardia lamblia

• Applicant: Jan Kosinski, Ph.D.
• Subject Area: Structural Biology ; Bioinformatics and Theoretical Biology ; Parasitology and Biology of Tropical Infectious Disease Pathogens ; Cell Biology
• Term: since 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 534243231

Giardia lamblia infects millions of people a year through contaminated food and water, causing diarrhoea, abdominal pain, fatigue, and weight loss. Drugs against Giardia work only for some infections and have substantial side effects - because humans and Giardia are both eukaryotes and share similar molecular machinery, developing drugs that target only the parasite has proven challenging. But there is a substantial molecular difference between humans and Giardia: endocytosis, the process by which eukaryotic cells take up nutrients and other molecules. Current knowledge is that, unlike other eukaryotes, endocytosis in Giardia lacks vesicles surrounded by a coat of clathrin proteins, which take substances from outside the cell and move deeper into the cell’s interior. Instead, Giardia uses so-called peripheral vacuoles: large, immobile organelles just below the cell membrane. To understand the finer details of how endocytosis works in Giardia, we imaged its cells using cutting-edge, high-resolution cryo-electron tomography (cryo-ET). To our surprise, we found vesicles covered by various coats including such that resemble clathrin-coated vesicles in size and appearance. Using AlphaFold2, an Artificial Intelligence-based program that predicts protein structures, we also found that endocytosis proteins in Giardia can actually form the complexes needed to assemble these vesicle coats. Both preliminary results suggest that Giardia’s endocytosis might, after all, involve clathrin-coated vesicles and that Giardia’s vesicular trafficking might be more conserved than previously proposed. This project will build on our surprising discoveries to resolve the true mechanism of endocytosis in Giardia. We aim to confirm which of these vesicles are indeed coated by clathrin and other coat types, trace the order of events observed in cryo-ET tomograms, and reveal the role of specific endocytosis proteins. To this end, we will combine time-resolved cryo-ET, structural modelling, and cell biology techniques including CRISPR/Cas9 genetic engineering. The molecular, structural, and mechanistic knowledge of Giardia’s endocytosis will provide a solid, evidence-based foundation for developing novel treatments against infection targeting specific features of Giardia’s endocytosis or exploiting it as a drug delivery pathway.

• DFG Programme: Research Grants