Natural variation of the drug-binding residues in eukaryotic ribosomes

Drugs that target eukaryotic ribosomes are becoming increasingly important as research tools and as potential therapies against cancer, fungi and other pathogenic eukaryotes. However, in the absence of comparative studies, we currently do not know how many eukaryotes possess ribosomal drug-binding sites identical to those in humans, and how many significantly differ from humans. Currently, this gap in knowledge is exacerbated by the presence of pseudogenes in eukaryotic genomes, making these comparative analyses challenging due to our inability to discriminate between genuine mutations, pseudogenes and sequencing artifacts. In this study, we resolve this problem by using a novel approach that leverages evolutionary relationships among species. Using this approach, we determine sequence variants for each of the 42 ribosomal drug-binding residues across 8,201 representative eukaryotes, tracing the evolutionary history of these variations from the emergence of eukaryotes 2 billion years ago to their subsequent divergence into distinct lineages. Unexpectedly, we find that yeasts and humans, commonly used as model eukaryotes to study ribosome/drug interactions, differ from most other eukaryotes due to the rRNA substitutions that mainly occur in animals and fungi but are absent in most other eukaryotes. Furthermore, we demonstrate that structural variants previously identified in pathogenic Leishmania and Plasmodium and viewed as idiosyncratic to a few eukaryotic species, are in fact shared by large clades of eukaryotes. Notably, some eukaryotic lineages exhibit ribosomal drug-binding sites that are more dissimilar to those of humans than humans are to bacteria. Overall, our study provides the most complete overview of ribosomal drug-binding sites evolution in eukaryotes, at the levels of single species, single residues, and single drugs, illuminating the eukaryotic lineages with structurally distinct ribosomal drug-binding sites compared to those of humans. These findings open new avenues for employing ribosome-targeting drugs as research tools and for the development of lineage-specific inhibitors against eukaryotic parasites.