|dc.description.abstract||Trypanosoma brucei differentiates into morphologically distinct forms as it inhabits two drastically different environments during its life cycle. In both stages, a single mitochondrion is required which differs greatly in the levels of activity during cell progression. The insect (procyclic) parasite has a very robust organelle that is active in energy processes such as the Krebs pathway and cytochrome mediated respiration while the mammalian (bloodstream) form has a repressed mitochondrion that is not active in these metabolic pathways. Despite these drastic differences, a diverse set of mitochondrial and nuclear encoded proteins is necessary for maintenance of this organelle in both developmental stages.
This work provides evidence of two contrasting mechanisms of protein diversity in T. brucei. The first part of this study further describes alternatively edited protein-1 (AEP-1), a polypeptide created from the post-transcriptional process of trypanosome mitochondrial RNA editing. Initially, I confirmed the presence of AEP-1 in procyclic and bloodstream form cells with a dual localization in their respective mitochondria. In both, AEP-1 resides in a nonionic detergent soluble complex associated with the inner mitochondrial membrane and a detergent resistant structure (tripartite attachment complex) that physically links the mitochondrial DNA (kinetoplast) to the basal bodies of the single flagellum of the cell. A previous study has revealed the importance of AEP-1 in genome integrity and characterizes this structural protein as a kinetoplast maintenance factor.
Sequencing of the soluble AEP-1 integral mitochondrial membrane complex (A-IMM) unsuspectingly uncovered moonlighting metabolic enzymes, another means of diversification, and revealed single proteins that have multiple functions within a cell. Here, I show that one of these proteins, the dihydrolipoamide succinyltransferase (E2) subunit of the α-ketoglutarate dehydrogenase, associates with the flagellum kinetoplast (kDNA) complex (FKC), which houses the tripartite attachment complex (TAC) and is important for kDNA distribution and mitochondrial biogenesis. Lastly, I demonstrate that depletion of a second enzyme from this Krebs pathway, the α-ketoglutarate dehydrogenase (E1) subunit, causes rapid swelling of an organelle involved in endo- and exocytosis of the cell, the flagellar pocket.||