DESCRIPTION (provided by the applicant): During the development of the asexual stage of the human malaria parasite, Plasmodium falciparum, the composition, structure and function of the host cell membrane is dramatically altered, including the ability to adhere to vascular endothelium. Crucial to these changes is the transport of parasite proteins, which become associated with the erythrocyte membrane. One of the most exciting problems in malaria cell biology is the mechanism by which intraerythocytic parasites target proteins to the RBC membrane, thus controlling its environment and contributing to the pathology of malaria. Protein targeting beyond the parasite plasma membrane (PPM) requires unique pathways, given the parasites intracellular location within a vacuolar membrane and the lack of organelles, secretory proteins and biosynthetic machinery in the host cell. It is not clear how these proteins cross the PVM or how they traverse the erythrocyte cytosol to reach their final destinations. It is our hypothesis that intraerythrocytic parasites export homologues of COPII, N-ethylmaleimide-sensitive factor (NSF) and other secretory proteins into the erythrocyte cytosol to generate a G-protein regulated secretory system that transports parasite proteins to the host cell membrane. Treatment of parasitized erythrocytes with aluminum fluoride (AIF) provided evidence of a G-protein mediated intraerythrocytic secretory pathway. In Aim 1, we will characterize the formation of intraerythrocytic transport vesicles, purify them from the erythrocyte of treated IRBC and determine their protein composition by LC/LC/MS/MS. Aim 2 is designed to investigate vesicle docking and fusion between intraerythrocytic secretory vesicles and the erythrocyte plasma membrane. Vesicle transport to the (parasite-derived) Maurer's clefts in the erythrocyte cytosol and the erythrocyte membrane appears to be actin-erythrocyte myosin mediated. Aim 3 will characterize the role of actin and myosin in vesicle formation and the transport of intraerythrocytic secretory vesicles to the RBC plasma membrane. The export and establishment of a specialized secretory system to a distant location is unusual in cell biology and points to the parasite 'transforming' the host cell into a secretory cell in order to meet the needs of the erythrocytic stage of infection. The identification of the key events in vesicle formation, transport, docking or fusion could lead to the identification of unique processes or molecules that might offer new drug targets.