Trypanosoma brucei: Differential requirement of membrane potential for import of proteins into mitochondria in two developmental stages

Trypanosome alternative oxidase (TAO) and the cytochrome oxidase (COX) are two developmentally regulated terminal oxidases of the mitochondrial electron transport chain in Trypanosoma brucei. Here, we have compared the import of TAO and cytochrome oxidase subunit IV (COIV), two stage-specific nuclear encoded mitochondrial proteins, into the bloodstream and procyclic form mitochondria of T. brucei to understand the import processes in two different developmental stages. Under in vitro conditions TAO and COIV were imported and processed into isolated mitochondria from both the bloodstream and procyclic forms. With mitochondria isolated from the procyclic form, the import of TAO and COIV was dependent on the mitochondrial inner membrane potential (Δψ) and required protein(s) on the outer membrane. Import of these proteins also depended on the presence of both internal and external ATP. However, import of TAO and COIV into isolated mitochondria from the bloodstream form was not inhibited after the mitochondrial Δψ was dissipated by valinomycin, CCCP, or valinomycin and oligomycin in combination. In contrast, import of these proteins into bloodstream mitochondria was abolished after the hydrolysis of ATP by apyrase or removal of the ATP and ATP-generating system, suggesting that import is dependent on the presence of external ATP. Together, these data suggest that nuclear encoded proteins such as TAO and COIV are imported in the mitochondria of the bloodstream and the procyclic forms via different mechanism. Differential import conditions of nuclear encoded mitochondrial proteins of T. brucei possibly help it to adapt to different life forms.