A high-resolution view of RNA endonuclease cleavage inBacillus subtilis

RNA endonucleases are the rate-limiting initiator of decay for many bacterial mRNAs. Subsequent decay by exonucleases typically obscures these cleavage events, and even for the well-studiedBacillus subtilis, few precise positions of cleavage have been mapped. Here we present an approach for precisely mapping positions of endoribonucleolytic activity transcriptome-wide. Through the detection of RNA ends in exonuclease-deficient cells, we map >103putative endonuclease cleavage sites, together with their dependence on RNase Y. Bioinformatic analysis reveals the targeting specificity of this enzyme, the mRNA interferase EndoA, as well as an unknown ribonuclease inB. subtilisthat trims the 5′ end of RNAs with both 5′-P and 5′-OH. Our results provide a high-resolution view of mRNA decay inB. subtilisand a generalizable approach for elucidating endoribonuclease specificityin vivo.