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Genome-wide libraries for protozoan pathogens drug target screening using yeast surface display

Rhiannon Heslop, Mengjin Gao, Andressa Brito Lira, Tamara Sternlieb, Mira Loock, Sahil Rao Sanghi, Igor Cestari
doi: https://doi.org/10.1101/2022.11.17.516844
Rhiannon Heslop
1Institute of Parasitology, McGill University, Ste Anne de Bellevue, QC H9X 3V9, Canada
3Faculté de Pharmacie de Tours, 31, Avenue Monge, 37200, Tours, France
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Mengjin Gao
1Institute of Parasitology, McGill University, Ste Anne de Bellevue, QC H9X 3V9, Canada
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Andressa Brito Lira
1Institute of Parasitology, McGill University, Ste Anne de Bellevue, QC H9X 3V9, Canada
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Tamara Sternlieb
1Institute of Parasitology, McGill University, Ste Anne de Bellevue, QC H9X 3V9, Canada
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Mira Loock
1Institute of Parasitology, McGill University, Ste Anne de Bellevue, QC H9X 3V9, Canada
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Sahil Rao Sanghi
1Institute of Parasitology, McGill University, Ste Anne de Bellevue, QC H9X 3V9, Canada
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Igor Cestari
1Institute of Parasitology, McGill University, Ste Anne de Bellevue, QC H9X 3V9, Canada
2Division of Experimental Medicine, McGill University, Montreal, QC, H4A 3J1, Canada
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  • For correspondence: igor.cestari@mcgill.ca
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Abstract

The lack of genetic tools to manipulate protozoan pathogens has limited the use of genome-wide approaches to identify drug or vaccine targets and understand these organisms’ biology. We have developed an efficient method to construct genome-wide libraries for yeast surface display (YSD) and developed a YSD fitness screen (YSD-FS) to identify drug targets. We show the robustness of our method by generating genome-wide libraries for Trypanosoma brucei, Trypanosoma cruzi, and Giardia lamblia parasites. Each library has a diversity of ∼105 to 106 clones, representing ∼6 to 30-fold of the parasite’s genome. Nanopore sequencing confirmed the libraries’ genome coverage with multiple clones for each parasite gene. Western blot and imaging analysis confirmed surface expression of the G. lamblia library proteins in yeast. Using the YSD-FS assay, we identified bonafide interactors of metronidazole, a drug used to treat protozoan and bacterial infections. We also found enrichment in nucleotide-binding domain sequences associated with yeast increased fitness to metronidazole, indicating that this drug might target multiple enzymes containing nucleotide-binding domains. The libraries are valuable biological resources for discovering drug or vaccine targets, ligand receptors, protein-protein interactions, and pathogen-host interactions. The library assembly approach can be applied to other organisms or expression systems, and the YSD-FS assay might help identify new drug targets in protozoan pathogens.

Competing Interest Statement

The authors have declared no competing interest.

Footnotes

  • https://github.com/cestari-lab/Libframe-tool

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted November 17, 2022.
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Genome-wide libraries for protozoan pathogens drug target screening using yeast surface display
Rhiannon Heslop, Mengjin Gao, Andressa Brito Lira, Tamara Sternlieb, Mira Loock, Sahil Rao Sanghi, Igor Cestari
bioRxiv 2022.11.17.516844; doi: https://doi.org/10.1101/2022.11.17.516844
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Genome-wide libraries for protozoan pathogens drug target screening using yeast surface display
Rhiannon Heslop, Mengjin Gao, Andressa Brito Lira, Tamara Sternlieb, Mira Loock, Sahil Rao Sanghi, Igor Cestari
bioRxiv 2022.11.17.516844; doi: https://doi.org/10.1101/2022.11.17.516844

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