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A CRISPR homing screen finds a chloroquine resistance transporter-like protein of the Plasmodium oocyst essential for mosquito transmission of malaria
Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
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2025 (English)In: Nature Communications, E-ISSN 2041-1723, Vol. 16, no 1, article id 3895Article in journal (Refereed) Published
Abstract [en]

Genetic screens with barcoded PlasmoGEM vectors have identified thousands of Plasmodium berghei gene functions in haploid blood stages, gametocytes and liver stages. However, the formation of diploid cells by fertilisation has hindered similar research on the parasites’ mosquito stages. In this study, we develop a scalable genetic system that uses barcoded gene targeting vectors equipped with a CRISPR-mediated homing mechanism to generate homozygous loss-of-function mutants after one parent introduces a modified allele into the zygote. To achieve this, we use vectors additionally expressing a target gene specific gRNA. When integrated into one of the parental alleles it directs Cas9 to the intact allele after fertilisation, leading to its disruption. This homing strategy is 90% effective at generating homozygous gene editing of a fluorescence-tagged reporter locus in the oocyst. A pilot screen identifies PBANKA_0916000 as a chloroquine resistance transporter-like protein (CRTL) essential for oocyst growth and sporogony, pointing to an unexpected importance for malaria transmission of the poorly understood digestive vacuole of the oocyst that contains hemozoin granules. Homing screens provide a method for the systematic discovery of malaria transmission genes whose first essential functions are after fertilisation in the bloodmeal, enabling their potential as targets for transmission-blocking interventions to be assessed.

Place, publisher, year, edition, pages
Springer Nature, 2025. Vol. 16, no 1, article id 3895
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Cell and Molecular Biology
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URN: urn:nbn:se:umu:diva-238485DOI: 10.1038/s41467-025-59099-1ISI: 001475587400017PubMedID: 40274854Scopus ID: 2-s2.0-105003447430OAI: oai:DiVA.org:umu-238485DiVA, id: diva2:1956717
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Knut and Alice Wallenberg FoundationAvailable from: 2025-05-07 Created: 2025-05-07 Last updated: 2025-05-07Bibliographically approved

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Balakrishnan, ArjunHunziker, MirjamTiwary, PujaPandey, VikashBillker, Oliver

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