Identification of a novel and ancestral machinery involved in mitochondrial membrane branching in Trypanosoma brucei

African trypanosomes are eukaryotic parasites that exist in two main replicative forms; the procyclic form in the midgut of the insect vector, the tsetse fly Glossina spp. and the bloodstream form responsible for diseases in humans and cattle. Unlike most other eukaryotes, where mitochondria continuously fuse and divide, trypanosome mitochondria form a single and continuously interconnected network that only divides during cytokinesis. The machineries governing mitochondrial remodeling and interconnection, however, remain largely unknown. We characterize a dynamin-related protein (DRP) from T. brucei (TbDBF, previously called TbMfnL) that depicts sequence similarities with Opa1 and Mfn, mammalian DRPs involved mitochondrial fusion. We showed that TbDBF has closely related homologues in several organisms that are devoid of Mfn and Opa1, such as eukaryotes from different phyla, prokaryotes and archaea. TbDBF is the first member of this new protein family to be functionally characterized. It localizes to the mitochondrial periphery and, upon overexpression, induces a strong increase in the interconnection and branching of mitochondrial filaments in a GTPase dependent manner. Its overexpression also promotes a major increase in cellular and mitochondrial volume and an increased consumption of the two major carbon sources used by the parasite (glucose and proline), as well as ethanolamine, a precursor of phosphatidyl-ethanolamine involved in membrane biogenesis and shaping. We propose that mitochondrial TbDBF is a component of an ancestral membrane remodeling machinery that contributes to the formation of intermitochondrial connections. ### Competing Interest Statement The authors have declared no competing interest.