To Catch a Killer: Analysis of linear plasmid and anticodon nuclease diversity across yeast species

Killer plasmids are a diverse group of virus-like elements (VLEs) present in four species of the Saccharomycotina, the budding yeasts. Killer plasmids encode “killer” toxins that cleave non-coding RNAs important for mRNA translation, such as tRNAs and rRNAs, allowing their hosts to kill competitors. These plasmids also encode immunity factors to their toxins to prevent the death of their hosts. Several species of the Saccharomycotina subphylum have non-standard genetic codes, where their CUG codon is translated as either serine or alanine instead of the canonical leucine. The faithfulness of this non-standard translation is due to translation of the CUG codon by novel tRNASerCAG and tRNAAlaCAG tRNAs. These have emerged through duplication events from pre-existing tRNA genes (either tRNASer or tRNAAla) followed by anticodon loop mutations. Their emergence is also accompanied by a selective loss of the ancestral tRNALeuCAG. Several species that continue to use the standard genetic code have lost the ancestral tRNALeuCAG genes, or have made major changes to tRNALeuCAG sequences, including large intron gains. We hypothesise that a killer toxin targeting the ancestral tRNALeuCAG selected for novel decoding of the CUG codon by the novel tRNASerCAG and tRNAAlaCAG tRNAs. In an effort to find evidence to support this hypothesis, I have found 45 novel toxin genes in the Saccharomycotina, both chromosomally- and VLE-encoded (Chapter 2). Expanding this search to wider fungi, I found 55 toxin genes chromosomally-encoded in the Pezizomycotina, the sister subphylum of the Saccharomycotina (Chapter 4). Finally, I provide the first evidence of linear dsDNA VLEs outside of the Saccharomycotina in Early Diverging Fungal lineages, Zoopagomycota and Mucoromycota, illuminating the origins of these elusive Saccharomycotina VLEs (Chapter 5).