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Publications (5 of 5) Show all publications
Russell, A. J. .., Sanderson, T., Bushell, E., Talman, A. M., Anar, B., Girling, G., . . . Billker, O. (2023). Regulators of male and female sexual development are critical for the transmission of a malaria parasite. Cell Host and Microbe, 31(2), 305-319.e10
Open this publication in new window or tab >>Regulators of male and female sexual development are critical for the transmission of a malaria parasite
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2023 (English)In: Cell Host and Microbe, ISSN 1931-3128, E-ISSN 1934-6069, Vol. 31, no 2, p. 305-319.e10Article in journal (Refereed) Published
Abstract [en]

Malaria transmission to mosquitoes requires a developmental switch in asexually dividing blood-stage parasites to sexual reproduction. In Plasmodium berghei, the transcription factor AP2-G is required and sufficient for this switch, but how a particular sex is determined in a haploid parasite remains unknown. Using a global screen of barcoded mutants, we here identify genes essential for the formation of either male or female sexual forms and validate their importance for transmission. High-resolution single-cell transcriptomics of ten mutant parasites portrays the developmental bifurcation and reveals a regulatory cascade of putative gene functions in the determination and subsequent differentiation of each sex. A male-determining gene with a LOTUS/OST-HTH domain as well as the protein interactors of a female-determining zinc-finger protein indicate that germ-granule-like ribonucleoprotein complexes complement transcriptional processes in the regulation of both male and female development of a malaria parasite.

Place, publisher, year, edition, pages
Cell Press, 2023
Keywords
development, differentiation, malaria, Plasmodium, sex determination, sex ratio, single cell analysis, transmission
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-212243 (URN)10.1016/j.chom.2022.12.011 (DOI)000964163800001 ()36634679 (PubMedID)2-s2.0-85147451944 (Scopus ID)
Funder
Wellcome trust, 206194/Z/17/ZKnut and Alice Wallenberg FoundationEU, European Research Council, 788516Wellcome trust, 083811Wellcome trust, 104111Wellcome trust, 107046Wellcome trust, 202600/Z/ 16/Z
Available from: 2023-07-20 Created: 2023-07-20 Last updated: 2023-07-20Bibliographically approved
Ishizaki, T., Hernandez, S., Paoletta, M. S., Sanderson, T. & Bushell, E. (2022). CRISPR/Cas9 and genetic screens in malaria parasites: small genomes, big impact. Biochemical Society Transactions, 50(3), 1069-1079
Open this publication in new window or tab >>CRISPR/Cas9 and genetic screens in malaria parasites: small genomes, big impact
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2022 (English)In: Biochemical Society Transactions, ISSN 0300-5127, E-ISSN 1470-8752, Vol. 50, no 3, p. 1069-1079Article, review/survey (Refereed) Published
Abstract [en]

The ∼30 Mb genomes of the Plasmodium parasites that cause malaria each encode ∼5000 genes, but the functions of the majority remain unknown. This is due to a paucity of functional annotation from sequence homology, which is compounded by low genetic tractability compared with many model organisms. In recent years technical breakthroughs have made forward and reverse genome-scale screens in Plasmodium possible. Furthermore, the adaptation of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-Associated protein 9 (CRISPR/Cas9) technology has dramatically improved gene editing efficiency at the single gene level. Here, we review the arrival of genetic screens in malaria parasites to analyse parasite gene function at a genome-scale and their impact on understanding parasite biology. CRISPR/Cas9 screens, which have revolutionised human and model organism research, have not yet been implemented in malaria parasites due to the need for more complex CRISPR/Cas9 gene targeting vector libraries. We therefore introduce the reader to CRISPR-based screens in the related apicomplexan Toxoplasma gondii and discuss how these approaches could be adapted to develop CRISPR/Cas9 based genome-scale genetic screens in malaria parasites. Moreover, since more than half of Plasmodium genes are required for normal asexual blood-stage reproduction, and cannot be targeted using knockout methods, we discuss how CRISPR/Cas9 could be used to scale up conditional gene knockdown approaches to systematically assign function to essential genes.

Place, publisher, year, edition, pages
Portland Press, 2022
Keywords
Plasmodium falciparum, biochemical techniques and resources, CRISPR, genetics, malaria
National Category
Cell and Molecular Biology Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-197999 (URN)10.1042/BST20210281 (DOI)000804375700001 ()35621119 (PubMedID)2-s2.0-85133214886 (Scopus ID)
Funder
Swedish Research Council, 2021-06602Knut and Alice Wallenberg Foundation, 2019.0178Wellcome trust, 210918/Z/18/Z
Available from: 2022-07-11 Created: 2022-07-11 Last updated: 2022-07-11Bibliographically approved
Marr, E. J., Milne, R. M., Anar, B., Girling, G., Schwach, F., Mooney, J. P., . . . Thompson, J. (2020). An enhanced toolkit for the generation of knockout and marker-free fluorescent Plasmodium chabaudi. Wellcome open research, 5, Article ID 71.
Open this publication in new window or tab >>An enhanced toolkit for the generation of knockout and marker-free fluorescent Plasmodium chabaudi
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2020 (English)In: Wellcome open research, ISSN 2398-502X, Vol. 5, article id 71Article in journal (Refereed) Published
Abstract [en]

The rodent parasite Plasmodium chabaudi is an important in vivo model of malaria. The ability to produce chronic infections makes it particularly useful for investigating the development of anti- Plasmodium immunity, as well as features associated with parasite virulence during both the acute and chronic phases of infection. P. chabaudi also undergoes asexual maturation (schizogony) and erythrocyte invasion in culture, so offers an experimentally-amenable in vivo to in vitro model for studying gene function and drug activity during parasite replication. To extend the usefulness of this model, we have further optimised transfection protocols and plasmids for P. chabaudi and generated stable, fluorescent lines that are free from drug-selectable marker genes. These mother-lines show the same infection dynamics as wild-type parasites throughout the lifecycle in mice and mosquitoes; furthermore, their virulence can be increased by serial blood passage and reset by mosquito transmission. We have also adapted the large-insert, linear PlasmoGEM vectors that have revolutionised the scale of experimental genetics in another rodent malaria parasite and used these to generate barcoded P. chabaudi gene-deletion and -tagging vectors for transfection in our fluorescent P. chabaudi mother-lines. This produces a tool-kit of P. chabaudi lines, vectors and transfection approaches that will be of broad utility to the research community.

Place, publisher, year, edition, pages
Wellcome open research, 2020
Keywords
PlasmoGem, Plasmodium chabaudi, Transfection, malaria
National Category
Microbiology in the medical area
Research subject
Microbiology
Identifiers
urn:nbn:se:umu:diva-172671 (URN)10.12688/wellcomeopenres.15587.1 (DOI)32500098 (PubMedID)2-s2.0-85086157248 (Scopus ID)
Available from: 2020-06-23 Created: 2020-06-23 Last updated: 2023-03-23Bibliographically approved
Stanway, R. R., Bushell, E., Chiappino-Pepe, A., Roques, M., Sanderson, T., Franke-Fayard, B., . . . Heussler, V. T. (2019). Genome-Scale Identification of Essential Metabolic Processes for Targeting the Plasmodium Liver Stage. Cell, 179(5), 1112-1128.e1-e15
Open this publication in new window or tab >>Genome-Scale Identification of Essential Metabolic Processes for Targeting the Plasmodium Liver Stage
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2019 (English)In: Cell, ISSN 0092-8674, E-ISSN 1097-4172, Vol. 179, no 5, p. 1112-1128.e1-e15Article in journal (Refereed) Published
Abstract [en]

Plasmodium gene functions in mosquito and liver stages remain poorly characterized due to limitations in the throughput of phenotyping at these stages. To fill this gap, we followed more than 1,300 barcoded P. berghei mutants through the life cycle. We discover 461 genes required for efficient parasite transmission to mosquitoes through the liver stage and back into the bloodstream of mice. We analyze the screen in the context of genomic, transcriptomic, and metabolomic data by building a thermodynamic model of P. berghei liver-stage metabolism, which shows a major reprogramming of parasite metabolism to achieve rapid growth in the liver. We identify seven metabolic subsystems that become essential at the liver stages compared with asexual blood stages: type II fatty acid synthesis and elongation (FAE), tricarboxylic acid, amino sugar, heme, lipoate, and shikimate metabolism. Selected predictions from the model are individually validated in single mutants to provide future targets for drug development.

Place, publisher, year, edition, pages
Elsevier, 2019
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-165826 (URN)10.1016/j.cell.2019.10.030 (DOI)000496914200010 ()31730853 (PubMedID)2-s2.0-85074715698 (Scopus ID)
Funder
Wellcome trust, 206194/Z/17/ZKnut and Alice Wallenberg Foundation
Note

Supplemental Figures

Available from: 2019-12-16 Created: 2019-12-16 Last updated: 2023-03-23Bibliographically approved
Hillier, C., Pardo, M., Yu, L., Bushell, E., Sanderson, T., Metcalf, T., . . . Choudhary, J. S. (2019). Landscape of the Plasmodium Interactome Reveals Both Conserved and Species-Specific Functionality. Cell Reports, 28(6), 1635-1647
Open this publication in new window or tab >>Landscape of the Plasmodium Interactome Reveals Both Conserved and Species-Specific Functionality
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2019 (English)In: Cell Reports, E-ISSN 2211-1247, Vol. 28, no 6, p. 1635-1647Article in journal (Refereed) Published
Abstract [en]

Malaria represents a major global health issue, and the identification of new intervention targets remains an urgent priority. This search is hampered by more than one-third of the genes of malaria-causing Plasmodium parasites being uncharacterized. We report a large-scale protein interaction network in Plasmodium schizonts, generated by combining blue native-polyacrylamide electrophoresis with quantitative mass spectrometry and machine learning. This integrative approach, spanning 3 species, identifies > 20,000 putative protein interactions, organized into 600 protein clusters. We validate selected interactions, assigning functions in chromatin regulation to previously unannotated proteins and suggesting a role for an EELM2 domain-containing protein and a putative microrchidia protein as mechanistic links between AP2-domain transcription factors and epigenetic regulation. Our interactome represents a high-confidence map of the native organization of core cellular processes in Plasmodium parasites. The network reveals putative functions for uncharacterized proteins, provides mechanistic and structural insight, and uncovers potential alternative therapeutic targets.

Place, publisher, year, edition, pages
Elsevier, 2019
Keywords
Plasmodium, blue native-PAGE, interactome, protein-protein interactions, interaction network, malaria, Plasmodium falciparum, Plasmodium berghei, Plasmodium knowlesi
National Category
Cell Biology
Identifiers
urn:nbn:se:umu:diva-162665 (URN)10.1016/j.celrep.2019.07.019 (DOI)000478978200023 ()31390575 (PubMedID)2-s2.0-85072149104 (Scopus ID)
Available from: 2019-09-04 Created: 2019-09-04 Last updated: 2024-01-17Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0003-2863-4112

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