FUNCTIONS AND ORIGINS OF MITOCHONDRION-RELATED ORGANELLES IN ANAEROBIC PROTISTS

Abstract

Across the diversity of life, organisms have evolved different strategies to thrive in hypoxic environments – microbial eukaryotes (protists) are no exception; protists that experience hypoxia often possess metabolically distinct mitochondria called mitochondrion-related organelles (MROs). Here, I focus on the biochemical adaptations of poorly-studied diverse anaerobic protists with an emphasis on the evolutionary histories of pyruvate, iron-sulfur cluster and respiratory chain metabolism in MROs. In the absence of oxygen, some organisms use the pyruvate formate lyase (PFL) system for the non-oxidative generation of acetyl-CoA. Through phylogenetic analyses, I showed that PFL is broadly, but patchily, distributed across the tree of eukaryotes. The monophyly of eukaryotes in these analyses suggest that the PFL pathway was first acquired by lateral gene transfer (LGT) into a eukaryotic lineage - from a firmicute bacterial lineage - and that it has since spread horizontally by more recent eukaryote-to-eukaryote transfer events. Biosynthesis of Fe-S clusters via the iron-sulfur cluster (ISC) system is a near-universally conserved feature of mitochondria and MROs. In contrast, some prokaryotic anaerobes synthesize and repair oxygen-damaged Fe-S clusters using a sulfur mobilization (SUF) system. Based on a transcriptomic survey, I reconstructed the MRO proteome of the protist Pygsuia biforma and found no evidence for the eukaryotic ISC system but instead identified a laterally acquired archaeabacterial-type SUF. Using immunofluorescence microscopy, I showed that SUF localizes to the MRO, representing the first reported case of a mitochondrial SUF system. In some anaerobes, the ubiquinone analog rhodoquinone (RQ) allows the respiratory complex II to function in reverse, generating succinate from fumarate. Using immunofluorescence microscopy, I demonstrated that a RQ biosynthesis protein (RQUA) localizes to Pygsuia MROs, suggesting the organelle participates in RQ biosynthesis. Phylogenetic analyses suggest that anaerobic eukaryotes acquired RQUA multiple times via LGT from distinct eukaryotic and bacterial donors. These studies suggest that crucial metabolic pathways localized to the MROs of anaerobic protists - and involved in their adaption to hypoxia - have been acquired by LGT. In some cases, these laterally acquired proteins were found to interface with ancestral mitochondrial proteins (e.g., acquired RQUA and the ancestrally mitochondrial complex II in Pygsuia) to create metabolic pathways of mosaic origins.