Recognition of a conserved class of RNA tetraloops by Saccharomyces cerevisiae RNase III

Ribonucleases III are double-stranded RNA (dsRNA) endonucleases required fo r the processing of a large number of prokaryotic and eukaryotic transcript s. Although the specificity of bacterial RNase III cleavage relies on antid eterminants in the dsRNA, the molecular basis of eukaryotic RNase III speci ficity is unknown. All substrates of yeast RNase ill (Rnt1p) are capped by terminal tetraloops showing the consensus AGNN and located within 13-16 bp to Rnt1p cleavage sites, We show that these tetraloops are essential for Rn t1p cleavage and that the distance to the tetraloop is the primary determin ant of cleavage site selection, The presence of AGNN tetraloops also enhanc es Rnt1p binding, as shown by surface plasmon resonance monitoring and modi fication interference studies, These results define a paradigm of RNA loops and show that yeast RNase III behaves as a helical RNA ruler that recogniz es these tetraloops and cleaves the dsRNA at a fixed distance to this RNA s tructure. These results also indicate that proteins belonging to the same c lass of RNA endonucleases require different structural elements for RNA cle avage.