INVESTIGATING A NOVEL CLADE OF ANAEROBIC MICROBIAL EUKARYOTES

Abstract

Metamonads are a diverse group of heterotrophic microbial eukaryotes adapted to living in hypoxic environments. All metamonads but one harbour metabolically altered ‘mitochondrion-related organelles’ (MROs) with reduced functions relative to aerobic mitochondria. To further investigate metamonad phylogeny, genome evolution and MRO diversity, we generated high-quality draft genomes, transcriptomes, and predicted proteomes for four novel free-living anaerobic flagellates: three species of the genus Skoliomonas and one Barthelona species. Phylogenomic analyses placed these organisms in a group we informally named the ‘BaSk’ (Barthelonids+Skoliomonads) clade, which emerges as a deeply branching sister group to the Fornicata, a metamonad phylum that includes parasitic and free-living flagellates. The sizes of the BaSk genomes vary significantly, ranging from 10.2 Mbp (Barthelona sp. PCE) to 33.5 Mbp (Skoliomonas sp. GEMRC). These differences are the result of the proliferation of transposable elements in some of these organisms, as well as differences in the sizes of gene families. Bioinformatic analyses of the gene models showed that these organisms have extremely reduced predicted MRO proteomes in comparison to other free-living metamonads. One of the isolates, Skoliomonas litria, potentially lacks a mitochondrial organelle altogether. If confirmed, this would be the first known example of complete mitochondrial loss in a free-living eukaryote. A systematic screen of these genomes for genes acquired by lateral transfer revealed several genes that likely supported their adaptation to hypoxia and reduction of their MROs. Notably, the lateral acquisition of a novel SUF-like minimal system (SMS) for Fe-S cluster biogenesis in a common ancestor of the BaSk group appears to have permitted the complete loss of the conserved mitochondrial iron-sulfur cluster (ISC) system in the skoliomonads. In parallel with the foregoing analyses, my colleagues and I developed a new pipeline for in silico prediction of mitochondrial or MRO localization of proteins called CoMR. CoMR consistently outperforms classic mitochondrial localization predictors, such as mitochondrial targeting sequence predictors, as well as new localization classification methods, such as internal targeting signal predictors. This thesis marks an important step towards understanding the evolution of the genomes of free-living anaerobic protists and introduces novel methods for studying their divergent mitochondria.