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Dual RNA-seq unveils noncoding RNA functions in host–pathogen interactions

Nature volume 529, pages 496501 (28 January 2016) | Download Citation

Abstract

Bacteria express many small RNAs for which the regulatory roles in pathogenesis have remained poorly understood due to a paucity of robust phenotypes in standard virulence assays. Here we use a generic ‘dual RNA-seq’ approach to profile RNA expression simultaneously in pathogen and host during Salmonella enterica serovar Typhimurium infection and reveal the molecular impact of bacterial riboregulators. We identify a PhoP-activated small RNA, PinT, which upon bacterial internalization temporally controls the expression of both invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity causes pervasive changes in coding and noncoding transcripts of the host. Interspecies correlation analysis links PinT to host cell JAK–STAT signalling, and we identify infection-specific alterations in multiple long noncoding RNAs. Our study provides a paradigm for a sensitive RNA-based analysis of intracellular bacterial pathogens and their hosts without physical separation, as well as a new discovery route for hidden functions of pathogen genes.

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Primary accessions

Gene Expression Omnibus

Data deposits

All RNA-seq data has been deposited in NCBI’s Gene Expression Omnibus under accession number GSE60144.

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Acknowledgements

We thank S. Gorski for help with the manuscript, C. Sharma and A. Eulalio for comments on the paper and L. Pfeuffer, B. Plaschke and V. McParland for technical assistance. We further thank S. Hoffmann for discussions about bioinformatic analyses, T. Rudel, K. Tedin, T. Meyer and C. Sharma for eukaryotic cell lines, V. Kozjak-Pavlovic for MitoTracker orange dye, T. Strowig for Ribo-Zero reagents, and W.-D. Hardt, M. Kolbe, H. Aiba and A. Eulalio for providing primary antibodies. A.J.W. was the recipient of an Elite Advancement Ph.D. stipend from the Universität Bayern e.V., Germany. L.B. is supported by a research fellowship from the Alexander von Humboldt Stiftung/Foundation. The Vogel laboratory received funding from the Bavarian BioSysNet program and BMBF (Bundesministerium für Bildung und Forschung) grant “Next-generation transcriptomics of bacterial infections”. This work was further supported by a donation from Baldwin Knauf to the Research Center for Infectious Diseases (ZINF). J.V. and P.F.S. acknowledge support by a joint BMBF grant eBio:RNAsys. This work used the European Grid Infrastructure (EGI) and funding by the EGI-Engage project (Horizon 2020) under grant number 654142.

Author information

Affiliations

  1. University of Würzburg, RNA Biology Group, Institute for Molecular Infection Biology, Josef-Schneider-Straße 2/D15, D-97080 Würzburg, Germany

    • Alexander J. Westermann
    • , Konrad U. Förstner
    • , Lars Barquist
    • , Yanjie Chao
    • , Leon N. Schulte
    •  & Jörg Vogel
  2. University of Würzburg, Core Unit Systems Medicine, Josef-Schneider-Straße 2/D15, D-97080 Würzburg, Germany

    • Konrad U. Förstner
  3. University of Leipzig, Department of Computer Science and Interdisciplinary Center for Bioinformatics, Härtelstraße 16-18, D-04107 Leipzig, Germany

    • Fabian Amman
    • , Lydia Müller
    •  & Peter F. Stadler
  4. University of Vienna, Theoretical Biochemistry Group, Institute for Theoretical Chemistry, Währinger Straße 17, A-1090 Vienna, Austria

    • Fabian Amman
    •  & Peter F. Stadler
  5. Max Planck Genome Centre Cologne, Max Planck Institute for Plant Breeding Research, Carl-von-Linné-Weg 10, D-50829 Cologne, Germany

    • Richard Reinhardt
  6. Max Planck Institute for Mathematics in the Sciences, Inselstraße 22, D-04103 Leipzig, Germany

    • Peter F. Stadler
  7. Santa Fe Institute, 1399 Hyde Park Rd, Santa Fe, New Mexico 87501, USA

    • Peter F. Stadler
  8. Research Centre for Infectious Diseases (ZINF), University of Würzburg, D-97070 Würzburg, Germany

    • Jörg Vogel

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Contributions

A.J.W., Y.C., L.N.S. and J.V. designed the research; A.J.W., Y.C. and L.N.S. performed the bench laboratory work; K.U.F., F.A., L.B., L.M. and P.F.S. conducted computational analyses; R.R. carried out part of the sequencing; A.J.W. and J.V. wrote the manuscript, which all co-authors commented on. J.V. supervised the project.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Jörg Vogel.

Extended data

Supplementary information

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  1. 1.

    Supplementary Figure 1

    This file contains gel source data.

Excel files

  1. 1.

    Supplementary Table 1

    This file contains information about the individual cDNA libraries sequenced, Salmonella pathway/regulon compositions, fold-changes in Salmonella gene expression upon PinT pulse-expression followed by sequencing, list of regulated host genes during the high-resolution comparative time-course Dual RNA-seq experiment, gene set and enriched GO-terms of the inter-species correlation analysis, strains, plasmids, oligonucleotides and antibodies used in this study.

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DOI

https://doi.org/10.1038/nature16547

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