Sequence features around cleavage sites are highly conserved among different species and a critical determinant for RNA cleavage position across eukaryotes.

RNA degradation is one of the critical steps for control of gene expression, and endonucleolytic cleavage-dependent RNA degradation is conserved among eukaryotes. Some cleavage sites are secondarily capped in the cytoplasm and identified using the Cap analysis of gene expression (CAGE) method. Although uncapped cleavage sites are widespread in eukaryotes, comparatively little information has been obtained about these sites using CAGE-based degradome analysis. Previously, we developed the truncated RNA-end sequencing (TREseq) method in plant species and used it to acquire comprehensive information about uncapped cleavage sites; we observed G-rich sequences near cleavage sites. However, it remains unclear whether this finding is general to other eukaryotes. In this study, we conducted TREseq analyses in fruit flies (Drosophila melanogaster) and budding yeast (Saccharomyces cerevisiae). The results revealed specific sequence features related to RNA cleavage in D. melanogaster and S. cerevisiae that were similar to sequence patterns in Arabidopsis thaliana. Although previous studies suggest that ribosome movements are important for determining cleavage position, feature selection using a random forest classifier showed that sequences around cleavage sites were major determinant for cleaved or uncleaved sites. Together, our results suggest that sequence features around cleavage sites are critical for determining cleavage position, and that sequence-specific endonucleolytic cleavage-dependent RNA degradation is highly conserved across eukaryotes.