Letter | Published:

NAD captureSeq indicates NAD as a bacterial cap for a subset of regulatory RNAs

Nature volume 519, pages 374377 (19 March 2015) | Download Citation

Abstract

A distinctive feature of prokaryotic gene expression is the absence of 5′-capped RNA. In eukaryotes, 5′,5′-triphosphate-linked 7-methylguanosine protects messenger RNA from degradation and modulates maturation, localization and translation1. Recently, the cofactor nicotinamide adenine dinucleotide (NAD) was reported as a covalent modification of bacterial RNA2. Given the central role of NAD in redox biochemistry, posttranslational protein modification and signalling3,4, its attachment to RNA indicates that there are unknown functions of RNA in these processes and undiscovered pathways in RNA metabolism and regulation. The unknown identity of NAD-modified RNAs has so far precluded functional analyses. Here we identify NAD-linked RNAs from bacteria by chemo-enzymatic capture and next-generation sequencing (NAD captureSeq). Among those identified, specific regulatory small RNAs (sRNAs) and sRNA-like 5′-terminal fragments of certain mRNAs are particularly abundant. Analogous to a eukaryotic cap, 5′-NAD modification is shown in vitro to stabilize RNA against 5′-processing by the RNA-pyrophosphohydrolase RppH5 and against endonucleolytic cleavage by ribonuclease (RNase) E6. The nudix phosphohydrolase NudC7 decaps NAD-RNA and thereby triggers RNase-E-mediated RNA decay, while being inactive against triphosphate-RNA. In vivo, 13% of the abundant sRNA RNAI is NAD-capped in the presence, and 26% in the absence, of functional NudC. To our knowledge, this is the first description of a cap-like structure and a decapping machinery in bacteria.

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Data deposits

NGS data have been deposited at http://www.geneprof.org under accession numbers gpXP_001108, gpXP_001123 and gpXP_001153.

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Acknowledgements

We thank the CellNetworks Deep Sequencing Core Facility for Solexa sequencing, M. Brunner for access to the light cycler, M. Helm for advice on RNA mass spectrometry and B. Luisi for the RNase E expression vector, as well as A. Krause, J. Becker, A. Samanta, M. Tesch, F. Siebert, L. Obenauer and other members of the Jäschke laboratory for help and discussions. H.C. was supported by a postdoctoral fellowship from the Alexander-von-Humboldt Foundation. M.-L.W. acknowledges a PhD fellowship from the Hartmut Hoffmann-Berling International Graduate School of Molecular & Cellular Biology. A.J. is supported by the Deutsche Forschungsgemeinschaft, SFB 623, the Federal Ministry of Education and Research (BMBF), and the Helmholtz Initiative on Synthetic Biology.

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    • Hana Cahová
    • , Marie-Luise Winz
    •  & Katharina Höfer

    These authors contributed equally to this work.

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  1. Institute of Pharmacy and Molecular Biotechnology (IPMB), Heidelberg University, 69120 Heidelberg, Germany

    • Hana Cahová
    • , Marie-Luise Winz
    • , Katharina Höfer
    • , Gabriele Nübel
    •  & Andres Jäschke

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Contributions

All authors designed the experiments, analysed and interpreted results and wrote the paper. H.C., M.-L.W., K.H. and G.N. performed the experiments.

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The authors declare no competing financial interests.

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Correspondence to Andres Jäschke.

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