Sleebs, Brad E3; Lopaticki, Sash3; Marapana, Danushka S3; O'Neill, Matthew T3; Rajasekaran, Pravin3; Gazdik, Michelle3; Günther, Svenja3; Whitehead, Lachlan W3; Lowes, Kym N3; Barfod, Lea6; Hviid, Lars6; Shaw, Philip J7; Hodder, Anthony N3; Smith, Brian J5; Cowman, Alan F3; Boddey, Justin A3
1 Centre for Medical Parasitology, Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, Københavns Universitet2 Department of Immunology and Microbiology, Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, Københavns Universitet3 The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia.4 HR-Unit, Faculty Service, Faculty of Humanities, Københavns Universitet5 Department of Chemistry, La Trobe University, Victoria, Australia.6 Centre for Medical Parasitology, Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, Københavns Universitet7 HR-Unit, Faculty Service, Faculty of Humanities, Københavns Universitet
The malaria parasite Plasmodium falciparum exports several hundred proteins into the infected erythrocyte that are involved in cellular remodeling and severe virulence. The export mechanism involves the Plasmodium export element (PEXEL), which is a cleavage site for the parasite protease, Plasmepsin V (PMV). The PMV gene is refractory to deletion, suggesting it is essential, but definitive proof is lacking. Here, we generated a PEXEL-mimetic inhibitor that potently blocks the activity of PMV isolated from P. falciparum and Plasmodium vivax. Assessment of PMV activity in P. falciparum revealed PEXEL cleavage occurs cotranslationaly, similar to signal peptidase. Treatment of P. falciparum-infected erythrocytes with the inhibitor caused dose-dependent inhibition of PEXEL processing as well as protein export, including impaired display of the major virulence adhesin, PfEMP1, on the erythrocyte surface, and cytoadherence. The inhibitor killed parasites at the trophozoite stage and knockdown of PMV enhanced sensitivity to the inhibitor, while overexpression of PMV increased resistance. This provides the first direct evidence that PMV activity is essential for protein export in Plasmodium spp. and for parasite survival in human erythrocytes and validates PMV as an antimalarial drug target.