The malaria digestive vacuole: elucidating its function in parasite physiology and development

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The malaria parasite Plasmodium falciparum is a single-celled microorganism with a complex life cycle that invades both host hepatocytes and red blood cells (RBCs). It spends a great proportion of its life cycle within the red blood cell (RBC), a cell comprised mostly of hemoglobin and lacking in nuclei and organelles. The invasion and subsequent modification of the RBC is a crucial step for the survival of this deadly parasite. To support the dynamic multiplication of malaria parasites, the parasite takes up large amounts of RBC cytosol into a specialized acidic organelle called the digestive vacuole (DV). This organelle breaks down the RBC cytosol’s primary constituent - hemoglobin - to provide itself with nutrients (i.e. amino acids) that are crucial for its growth. The DV has been thought to have similarities to both tonoplasts, an acidic intracellular vacuole of plant cells, and lysosomes of mammalian cells. However, the absence of the typical lysosomal acid phosphatase and glycosidase indicates that the DV of Plasmodium is a specialized organelle that most likely evolved to efficiently degrade hemoglobin. Consequently, the parasite’s DV carries out a variety of specialized and critical functions to ensure the survival of the parasite, including hemoglobin degradation, detoxification of oxygen radicals, ion homeostasis, and nutrient and/or solute transport across its membrane. The two most prominent and essential transporters (the multi-drug transporter PfMDR1 and the chloroquine resistance transporter PfCRT) situated on the DV membrane are yet to be fully characterized. The functional role of these transporters, as well as the substrate(s) they transport, remains obscure. It is clear, however, that they did not evolve within the parasite simply to efflux drugs. The underlying biology of this complex organelle is incomplete and poorly understood, possibly due to past experiments that mostly used fixed samples. Therefore, the objective of this research program is the development of an in-depth understanding of the molecular and cellular processes of the parasite’s digestive vacuole using modern imaging techniques tailored to P. falciparum and available in our lab. With our previous NSERC proposal, we set out to better characterize the PfMDR1 transporter. We accomplished an important objective of this proposal in that we were able to quantify the kinetics of the PfMDR1 transporter in live parasites using a reverse imaging assay, a technique that one of the journal reviewers termed “ingenious”. The present proposal will keep our focus on the quantification of various important processes occurring in the DV. We will continue to use live cell imaging techniques to get a better understanding of parasite dynamics in situ. This is a unique approach that is used in only a limited number of labs world-wide and will allow us to better understand these processes in real time within the live parasite.