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  • 1. Hillier, Charles
    et al.
    Pardo, Mercedes
    Yu, Lu
    Bushell, Ellen
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Sanderson, Theo
    Metcalf, Tom
    Herd, Colin
    Anar, Burcu
    Rayner, Julian C.
    Billker, Oliver
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Choudhary, Jyoti S.
    Landscape of the Plasmodium Interactome Reveals Both Conserved and Species-Specific Functionality2019Inngår i: Cell Reports, E-ISSN 2211-1247, Vol. 28, nr 6, s. 1635-1647Artikkel i tidsskrift (Fagfellevurdert)
    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.

    Fulltekst (pdf)
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  • 2.
    Ishizaki, Takahiro
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Hernandez, Sophia
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Paoletta, Martina S.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Instituto de Agrobiotecnología y Biología Molecular (IABIMO), Argentina.
    Sanderson, Theo
    Francis Crick Institute London, United Kingdom.
    Bushell, Ellen
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    CRISPR/Cas9 and genetic screens in malaria parasites: small genomes, big impact2022Inngår i: Biochemical Society Transactions, ISSN 0300-5127, E-ISSN 1470-8752, Vol. 50, nr 3, s. 1069-1079Artikkel, forskningsoversikt (Fagfellevurdert)
    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.

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  • 3. Marr, Edward J
    et al.
    Milne, Rachel M
    Anar, Burcu
    Girling, Gareth
    Schwach, Frank
    Mooney, Jason P
    Nahrendorf, Wiebke
    Spence, Philip J
    Cunningham, Deirdre
    Baker, David A
    Langhorne, Jean
    Rayner, Julian C
    Billker, Oliver
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Parasites and Microbes, Wellcome Sanger Institute, Wellcome Genome Campus, Cambridge, UK.
    Bushell, Ellen S.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Parasites and Microbes, Wellcome Sanger Institute, Wellcome Genome Campus, Cambridge, UK.
    Thompson, Joanne
    An enhanced toolkit for the generation of knockout and marker-free fluorescent Plasmodium chabaudi2020Inngår i: Wellcome open research, ISSN 2398-502X, Vol. 5, artikkel-id 71Artikkel i tidsskrift (Fagfellevurdert)
    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.

  • 4.
    Russell, Andrew J.C.
    et al.
    Wellcome Sanger Institute, Hinxton, United Kingdom.
    Sanderson, Theo
    Wellcome Sanger Institute, Hinxton, United Kingdom; Francis Crick Institute, 1 Midland Road, London, United Kingdom.
    Bushell, Ellen
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Wellcome Sanger Institute, Hinxton, United Kingdom.
    Talman, Arthur M.
    Wellcome Sanger Institute, Hinxton, United Kingdom; MIVEGEC, University of Montpellier, IRD, CNRS, Montpellier, France.
    Anar, Burcu
    Wellcome Sanger Institute, Hinxton, United Kingdom.
    Girling, Gareth
    Wellcome Sanger Institute, Hinxton, United Kingdom.
    Hunziker, Mirjam
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Kent, Robyn S.
    Department of Microbiology and Molecular Genetics, University of Vermont Larner College of Medicine, VT, Burlington, United States.
    Martin, Julie S.
    Wellcome Centre for Integrative Parasitology, Institute of Infection, Immunity, and Inflammation, University of Glasgow, Glasgow, United Kingdom.
    Metcalf, Tom
    Wellcome Sanger Institute, Hinxton, United Kingdom.
    Montandon, Ruddy
    Wellcome Sanger Institute, Hinxton, United Kingdom.
    Pandey, Vikash
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Pardo, Mercedes
    The Institute of Cancer Research, London, United Kingdom.
    Roberts, A. Brett
    Wellcome Centre for Integrative Parasitology, Institute of Infection, Immunity, and Inflammation, University of Glasgow, Glasgow, United Kingdom.
    Sayers, Claire
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Schwach, Frank
    Wellcome Sanger Institute, Hinxton, United Kingdom.
    Choudhary, Jyoti S.
    The Institute of Cancer Research, London, United Kingdom.
    Rayner, Julian C.
    Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom.
    Voet, Thierry
    Department of Human Genetics, University of Leuven, KU Leuven, Leuven, Belgium; KU Leuven Institute for Single Cell Omics, LISCO, KU Leuven, Leuven, Belgium.
    Modrzynska, Katarzyna K.
    Wellcome Centre for Integrative Parasitology, Institute of Infection, Immunity, and Inflammation, University of Glasgow, Glasgow, United Kingdom.
    Waters, Andrew P.
    Wellcome Centre for Integrative Parasitology, Institute of Infection, Immunity, and Inflammation, University of Glasgow, Glasgow, United Kingdom.
    Lawniczak, Mara K.N.
    Wellcome Sanger Institute, Hinxton, United Kingdom.
    Billker, Oliver
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Regulators of male and female sexual development are critical for the transmission of a malaria parasite2023Inngår i: Cell Host and Microbe, ISSN 1931-3128, E-ISSN 1934-6069, Vol. 31, nr 2, s. 305-319.e10Artikkel i tidsskrift (Fagfellevurdert)
    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.

    Fulltekst (pdf)
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  • 5. Stanway, Rebecca R.
    et al.
    Bushell, Ellen
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Chiappino-Pepe, Anush
    Roques, Magali
    Sanderson, Theo
    Franke-Fayard, Blandine
    Caldelari, Reto
    Golomingi, Murielle
    Nyonda, Mary
    Pandey, Vikash
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Laboratory of Computational Systems Biotechnology, École Polytechnique Fédérale de Lausanne, EPFL, Lausanne 1015, Switzerland.
    Schwach, Frank
    Chevalley, Séverine
    Ramesar, Jai
    Metcalf, Tom
    Herd, Colin
    Burda, Paul-Christian
    Rayner, Julian C.
    Soldati-Favre, Dominique
    Janse, Chris J.
    Hatzimanikatis, Vassily
    Billker, Oliver
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet). Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge CB10 1SA, UK.
    Heussler, Volker T.
    Genome-Scale Identification of Essential Metabolic Processes for Targeting the Plasmodium Liver Stage2019Inngår i: Cell, ISSN 0092-8674, E-ISSN 1097-4172, Vol. 179, nr 5, s. 1112-1128.e1-e15Artikkel i tidsskrift (Fagfellevurdert)
    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.

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