A high-resolution view of RNA endonuclease cleavage in Bacillus subtilis

Taggart, J.; Lalanne, J.-B.; Durand, S.; Braun, F.; Condon, C.; Li, G.-W.

doi:10.1101/2023.03.12.532304

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bioRxiv · 10.1101/2023.03.12.532304

A high-resolution view of RNA endonuclease cleavage in Bacillus subtilis

Taggart, J.·Lalanne, J.-B.·Durand, S.·Braun, F.·Condon, C.·Li, G.-W.

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Abstract

RNA endonucleases are the rate-limiting initiator of decay for many bacterial mRNAs. However, the positions of cleavage and their sequence determinants remain elusive even for the well-studied Bacillus subtilis. Here we present two complementary approaches - transcriptome-wide mapping of endoribonucleolytic activity and deep mutational scanning of RNA cleavage sites - that reveal distinct rules governing the specificity among B. subtilis endoribonucleases. Detection of RNA terminal nucleotides in both 5'- and 3'-exonuclease-deficient cells revealed >103 putative endonucleolytic cleavage sites with single-nucleotide resolution. We found a surprisingly weak consensus for RNase Y targets, a contrastingly strong primary sequence motif for EndoA targets, and long-range intramolecular secondary structures for RNase III targets. Deep mutational analysis of RNase Y cleavage sites showed that the specificity is governed by many disjointed sequence features, each with mild contributions. Our results highlight the delocalized nature of mRNA stability determinants and provide a strategy for elucidating endoribonuclease specificity in vivo.

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Taggart, J., Lalanne, J.-B., Durand, S., Braun, F., Condon, C., Li, G.-W.. 2023-03-13. A high-resolution view of RNA endonuclease cleavage in Bacillus subtilis. https://doi.org/10.1101/2023.03.12.532304

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