title: Conditional Degradation of Plasmodium Calcineurin Reveals Functions in Parasite Colonization of both Host and Vector.
abstract: Functional analysis of essential genes in the malarial parasite, Plasmodium, is hindered by lack of efficient strategies for conditional protein regulation. We report the development of a rapid, specific, and inducible chemical-genetic tool in the rodent malaria parasite, P. berghei, in which endogenous proteins engineered to contain the auxin-inducible degron (AID) are selectively degraded upon adding auxin. Application of AID to the calcium-regulated protein phosphatase, calcineurin, revealed functions in host and vector stages of parasite development. Whereas depletion of calcineurin in late-stage schizonts demonstrated its critical role in erythrocyte attachment and invasion in vivo, stage-specific depletion uncovered roles in gamete development, fertilization, and ookinete-to-oocyst and sporozoite-to-liver stage transitions. Furthermore, AID technology facilitated concurrent generation and phenotyping of transgenic lines, allowing multiple lines to be assessed simultaneously with significant reductions in animal use. This study highlights the broad applicability of AID for functional analysis of proteins across the Plasmodium life cycle.
authors.affiliation: Wellcome Trust Centre for Molecular Parasitology, Institute of Infection, Immunity and Inflammation, College of Medical Veterinary & Life Sciences, University of Glasgow, Glasgow, G12 8TA, UK. Electronic address: Nisha.Philip@glasgow.ac.uk.
authors.forename: Nisha
authors.initials: N
authors.lastname: Philip
authors.affiliation: Wellcome Trust Centre for Molecular Parasitology, Institute of Infection, Immunity and Inflammation, College of Medical Veterinary & Life Sciences, University of Glasgow, Glasgow, G12 8TA, UK. Electronic address: Andy.Waters@glasgow.ac.uk.
authors.forename: Andrew P
authors.initials: AP
authors.lastname: Waters
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D019703:Calcineurin / Q000235:genetics / Q000378:metabolism*; D009033:Culicidae; D006790:Host-Parasite Interactions; D007210:Indoleacetic Acids / Q000378:metabolism*; D051379:Mice; D010962:Plasmodium berghei / Q000254:growth & development / Q000502:physiology*; D059748:Proteolysis / Q000187:drug effects*; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D007210:Indoleacetic Acids; D015800:Protozoan Proteins; D019703:Calcineurin