The Plasmodium falciparum protein VCAP1 controls Maurer’s cleft morphology, knob architecture and Pf EMP1 trafficking

Abstract The malaria parasite, Plasmodium falciparum , traffics the virulence protein, P. falciparum erythrocyte membrane protein 1 ( Pf EMP1) to the surface of infected red blood cells (RBCs) via membranous organelles, known as the Maurer’s clefts. We developed a method for efficient enrichment of Maurer’s clefts and profiled the protein composition of this trafficking organelle. We identified 13 previously uncharacterised or poorly characterised Maurer’s cleft proteins. We generated transfectants expressing GFP-fusions of 7 proteins and confirmed their Maurer’s cleft location. Using co-immunoprecipitation and mass spectrometry we have generated a protein interaction map of proteins at the Maurer’s clefts. We identified two key clusters that may function in the loading and unloading of Pf EMP1 into and out of the Maurer’s clefts. We focus on a putative Pf EMP1 loading complex that includes the newly characterised virulence complex assembly protein 1 (VCAP1). Disruption of VCAP1 causes Maurer’s cleft fragmentation, aberrant knobs, ablation of Pf EMP1 surface expression and loss of the Pf EMP1 directed adhesion. ΔVCAP1 parasite lines have a growth advantage compared to wildtype parasites; and the infected RBCs are more deformable and more osmotically fragile. Importance The trafficking of the virulence antigen Pf EMP1 and its presentation at the knob structures at the surface of parasite infected RBCs is central to severe adhesion related pathologies such as cerebral and placental malaria. This work adds to our understanding of how PfEMP1 is trafficked to the RBC membrane by defining the protein-protein interaction networks that function at the Maurer’s clefts controlling PfEMP1 loading and unloading. This work adds significantly to our understanding of virulence protein trafficking and will provide crucial knowledge that will be required to determine the mechanisms underpinning parasite driven host cell remodelling, parasite survival within the host and virulence mechanisms.