Abstract
BACKGROUND: In the budding yeast Saccharomyces cerevisiae, the widespread adoption of ribosome profiling technology has allowed the discovery of evidence of transcription and translation for thousands of small proteins or microproteins whose importance was once disregarded. Both conserved and evolutionarily short-lived microproteins have demonstrated relevant involvement in biological functions. However, sequences exist in a broad spectrum of conservation. Here, we tested whether these small proteins in yeast detected by ribosome profiling technology have different properties across their levels of conservation, and how do these properties compare with the canonical small protein-coding sequences. RESULTS: Here, we applied a phylostratigraphic approach to peptides encoded by small open reading frames. We compared 20,023 ribo-seq-detected small peptides against annotated small proteins belonging to reference annotations on the basis of their respective conservation patterns. We identified 1134 unannotated microproteins that, despite their difficulty in being detected by methods other than ribosome profiling, display hallmarks of functionality such as conservation across many taxonomical levels and signals of purifying selection not dissimilar to those of canonical proteins of comparable length. Sequences that initially did not show evidence of belonging to any gene family were found to possess signals of homology traceable mostly at genus level when compared against noncoding regions and using TBLASTN, but also, to a lesser extent, to species belonging to the phyla Basidiomycota and Microsporidia. In addition, we show an analysis of the mutations behind the origin of small open reading frames exclusive to S. cerevisiae and identified changes in the initiation codon as the most common group of mutations when compared to Saccharomyces paradoxus, the closest species to S. cerevisiae. CONCLUSIONS: Our work, by presenting robust analysis of the extended landscape of small proteins in yeast, suggests that small conserved sequences, either canonical or not, possess a shared evolutionary trajectory, as demonstrated by their properties. These results shed some light into the evolutionary processes behind the extended landscape of small proteins in yeast.