Functional characterization of phospholipases in the malaria parasite Plasmodium falciparum

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Plasmodium parasites, the causative agents of malaria, infect hepatocytes and red blood cells (RBCs). Within these cells, parasites are surrounded by a parasitophorous vacuole membrane (PVM). Consequently, for their release from host cells and to propagate their infection, parasites have to disrupt the PVM and the host cell membrane. How this membrane rupture occurs on a molecular level and how it is controlled is only partially understood and only a very limited number of parasite proteins involved in this process have been identified so far. We previously showed that a parasite phospholipase mediates PVM rupture during parasite egress from hepatocytes. Whether parasite phospholipases are similarly involved in parasite release from RBCs is not known and most of the Plasmodium phospholipases are not characterized. In this project, it is thus proposed to functionally analyze the role of phospholipases of the human malaria parasite Plasmodium falciparum during its development within red blood cells. First, we will probe into the redundancy of those enzymes that show expression in asexual blood stage parasites by targeted gene disruption. Essential genes, which cannot be deleted using conventional knockout techniques, will further be analyzed using conditional systems, their loss-of-function phenotypes analyzed and their cellular localization elucidated. In order to reveal a potential interplay and synergistic functions between different phospholipases, double knockout parasites targeting a subset of enzymes will be generated and analyzed. Finally, we aim to investigate, whether parasite phospholipases are activated by proteolytic cleavage of the egress-associated subtilisin-like serine protease SUB1. For this, the functional importance of predicted SUB1 cleavage sites identified in several putative phospholipases will be tested. In conclusion, our work will increase our knowledge of the molecular machinery leading to parasite release from host cells and might lead to the identification of novel targets for parasite-specific intervention strategies.