The mitochondrial calcium uniporter of trypanosomes

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DESCRIPTION (provided by applicant): The T. brucei group of parasites is the causative agent of sleeping sickness or African trypanosomiasis. According to the World Health Organization over 60 million people in sub-Saharan Africa are at risk of infection with an incidence of thousands of cases per year. African trypanosomiasis has been reemerging since the 1970s, and chemotherapy remains unsatisfactory especially for advanced cases. T. brucei is, in addition, the trypanosome most amenable to molecular and genetic experimentation, and for which powerful genetic tools have been developed. Under physiological conditions mitochondrial Ca2+ uptake takes place by the uniport mechanism driven electrophoretically by the negative-inside membrane potential without direct coupling to ATP hydrolysis or transport of other ions. This MCU was found more than 50 years ago (1961- 1962) in rodent mitochondria and the biophysical properties of this Ca2+-selective channel were extensively characterized since then. For many years after the discovery of the MCU in mammalian mitochondria, it was thought that less complex life forms such as plants, insects and other invertebrates, or unicellular organisms such as yeast, lacked a specific mitochondrial calcium uptake pathway. This was until we reported, in 1989, that epimastigotes of Trypanosoma cruzi, the etiologic agent of Chagas disease, possesses a MCU with characteristics similar to those described in mammalian mitochondria: electrogenic transport, sensitivity to ruthenium red, and low affinity for the cation. The evidence of the presence of a MCU in trypanosomes but its absence in yeast was the key to the discovery of the molecular identity of MCU. Mammalian mitochondria have a central role in intracellular Ca2+ homeostasis, and it is well established that intramitochondrial Ca2+ concentration can reach tens or hundreds micromolar values upon cytosolic Ca2+ rises of a few micromolar. This is because mitochondria are exposed to microdomains of high Ca2+ concentration in proximity to sites of Ca2+ release at the endoplasmic reticulum, or to Ca2+ channels at the plasma membrane. This Ca2+ uptake is important for shaping the amplitude and spatio-temporal patterns of cytosolic Ca2+ increases and for regulating the activity of three intramitochondrial dehydrogenases that result in ATP generation, as well as the activity of the ATP synthase. Excessive Ca2+ uptake, however, favors the formation of the "permeability transition pore" leading to the release of proapoptotic factors in the cytosol and cell death. Interestingly, T. brucei mitochondrion is also exposed to high Ca2+ levels in proximity to acidocalcisomes, acidic calcium storage compartments that we discovered in T. brucei in 1994, and that contain the inositol 1,4,5- triphosphate receptor (IP3R) for Ca2+ release. Our hypothesis is that T. brucei could be used to investigate the essentiality of the MCU and mitochondrial Ca2+ uptake, the relation between mitochondria and acidocalcisomes, and the presence of other components of the mitochondrial Ca2+ uptake complex that have been postulated to exist.