title: Localisation-based imaging of malarial antigens during erythrocyte entry reaffirms a role for AMA1 but not MTRAP in invasion.
abstract: Microscopy-based localisation of proteins during malaria parasite (Plasmodium) invasion of the erythrocyte is widely used for tentative assignment of protein function. To date, however, imaging has been limited by the rarity of invasion events and the poor resolution available, given the micron size of the parasite, which leads to a lack of quantitative measures for definitive localisation. Here, using computational image analysis we have attempted to assign relative protein localisation during invasion using wide-field deconvolution microscopy. By incorporating three-dimensional information we present a detailed assessment of known parasite effectors predicted to function during entry but as yet untested or for which data are equivocal. Our method, termed longitudinal intensity profiling, resolves confusion surrounding the localisation of apical membrane antigen 1 (AMA1) at the merozoite-erythrocyte junction and predicts that the merozoite thrombospondin-related anonymous protein (MTRAP) is unlikely to play a direct role in the mechanics of entry, an observation supported with additional biochemical evidence. This approach sets a benchmark for imaging of complex micron-scale events and cautions against simplistic interpretations of small numbers of representative images for the assignment of protein function or prioritisation of candidates as therapeutic targets.
authors.affiliation: The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, 3052, Australia Department of Medical Biology, University of Melbourne, Victoria, 3050, Melbourne, Australia.
authors.forename: David T
authors.initials: DT
authors.lastname: Riglar
authors.affiliation: The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, 3052, Australia Department of Medical Biology, University of Melbourne, Victoria, 3050, Melbourne, Australia.
authors.forename: Lachlan
authors.initials: L
authors.lastname: Whitehead
authors.affiliation: The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, 3052, Australia Department of Medical Biology, University of Melbourne, Victoria, 3050, Melbourne, Australia.
authors.forename: Alan F
authors.initials: AF
authors.lastname: Cowman
authors.affiliation: The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, 3052, Australia Department of Medical Biology, University of Melbourne, Victoria, 3050, Melbourne, Australia.
authors.forename: Kelly L
authors.initials: KL
authors.lastname: Rogers
authors.affiliation: Department of Life Sciences, Imperial College, London SW7 2AZ, UK jake.baum@imperial.ac.uk.
authors.forename: Jake
authors.initials: J
authors.lastname: Baum
mesh_terms: D000199:Actins / Q000378:metabolism; D000913:Antibodies, Protozoan / Q000378:metabolism; D000918:Antibody Specificity; D000953:Antigens, Protozoan / Q000378:metabolism*; D000939:Epitopes / Q000378:metabolism; D004912:Erythrocytes / Q000469:parasitology*; D005634:Fructose-Bisphosphate Aldolase / Q000378:metabolism; D006801:Humans; D021621:Imaging, Three-Dimensional; D052940:Merozoites / Q000378:metabolism; D008954:Models, Biological; D010963:Plasmodium falciparum / Q000276:immunology* / Q000502:physiology*; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D015800:Protozoan Proteins / Q000378:metabolism*; D019108:Tight Junctions / Q000378:metabolism
chemical_list: D000199:Actins; D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D000939:Epitopes; D015800:Protozoan Proteins; D005634:Fructose-Bisphosphate Aldolase
keywords: Deconvolution; Erythrocyte invasion; Merozoite; Plasmodium falciparum; Tight junction