Dynamic interactions between the RNA chaperone Hfq, small regulatory RNAs and mRNAs in live bacterial cells

Abstract

RNA-binding proteins play myriad roles in regulating RNAs and RNA-mediated functions. In bacteria, the RNA chaperone Hfq is an important post-transcriptional gene regulator. Using live-cell super-resolution imaging, we can distinguish Hfq binding to different sizes of cellular RNAs. We demonstrate that under normal growth conditions, Hfq exhibits widespread mRNA-binding activity, with the distal face of Hfq contributing mostly to the mRNA binding in vivo. In addition, sRNAs can either co-occupy Hfq with the mRNA as a ternary complex, or displace the mRNA from Hfq in a binding face-dependent manner, suggesting mechanisms through which sRNAs rapidly access Hfq to induce sRNA-mediated gene regulation. Finally, our data suggest that binding of Hfq to certain mRNAs through its distal face can recruit RNase E to promote turnover of these mRNAs in an sRNA-independent manner, and such regulatory function of Hfq can be decoyed by sRNA competitors that bind strongly at the distal face.

Data availability

All the numeric data for each plot/graph and fitting results are provided in Supplementary file 1 or as source data. The MATLAB scripts for analysis are provided as source code.

Article and author information

Author details

  1. Seongjin Park

    Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Karine Prévost

    RNA Group, Department of Biochemistry, University of Sherbrooke, Sherbrooke, Canada
    Competing interests
    The authors declare that no competing interests exist.
  3. Emily M Heideman

    Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Marie-Claude Carrier

    RNA Group, Department of Biochemistry, University of Sherbrooke, Sherbrooke, Canada
    Competing interests
    The authors declare that no competing interests exist.
  5. Muhammad S Azam

    Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Matthew A Reyer

    Institute for Biophysical Dynamics, University of Chicago, Chicago, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Wei Liu

    Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Eric Massé

    RNA Group, Department of Biochemistry, University of Sherbrooke, Sherbrooke, Canada
    For correspondence
    eric.masse@usherbrooke.ca
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5253-1415
  9. Jingyi Fei

    Department of Biochemistry and Molecular Biology, Institute for Biophysical Dynamics, University of Chicago, Chicago, United States
    For correspondence
    jingyifei@uchicago.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9775-3820

Funding

National Institutes of Health (1DP2GM128185-01)

  • Jingyi Fei

Searle Scholars Program

  • Jingyi Fei

National Institutes of Health (R01 GM092830-06A1)

  • Eric Massé

Canadian Institutes of Health Research (MOP69005)

  • Eric Massé

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2021, Park et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 3,855
    views
  • 514
    downloads
  • 31
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Seongjin Park
  2. Karine Prévost
  3. Emily M Heideman
  4. Marie-Claude Carrier
  5. Muhammad S Azam
  6. Matthew A Reyer
  7. Wei Liu
  8. Eric Massé
  9. Jingyi Fei
(2021)
Dynamic interactions between the RNA chaperone Hfq, small regulatory RNAs and mRNAs in live bacterial cells
eLife 10:e64207.
https://doi.org/10.7554/eLife.64207

Share this article

https://doi.org/10.7554/eLife.64207

Further reading

    1. Biochemistry and Chemical Biology
    2. Stem Cells and Regenerative Medicine
    Alejandro J Brenes, Eva Griesser ... Angus I Lamond
    Research Article

    Human induced pluripotent stem cells (hiPSCs) have great potential to be used as alternatives to embryonic stem cells (hESCs) in regenerative medicine and disease modelling. In this study, we characterise the proteomes of multiple hiPSC and hESC lines derived from independent donors and find that while they express a near-identical set of proteins, they show consistent quantitative differences in the abundance of a subset of proteins. hiPSCs have increased total protein content, while maintaining a comparable cell cycle profile to hESCs, with increased abundance of cytoplasmic and mitochondrial proteins required to sustain high growth rates, including nutrient transporters and metabolic proteins. Prominent changes detected in proteins involved in mitochondrial metabolism correlated with enhanced mitochondrial potential, shown using high-resolution respirometry. hiPSCs also produced higher levels of secreted proteins, including growth factors and proteins involved in the inhibition of the immune system. The data indicate that reprogramming of fibroblasts to hiPSCs produces important differences in cytoplasmic and mitochondrial proteins compared to hESCs, with consequences affecting growth and metabolism. This study improves our understanding of the molecular differences between hiPSCs and hESCs, with implications for potential risks and benefits for their use in future disease modelling and therapeutic applications.

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Jie Luo, Jeff Ranish
    Tools and Resources

    Dynamic conformational and structural changes in proteins and protein complexes play a central and ubiquitous role in the regulation of protein function, yet it is very challenging to study these changes, especially for large protein complexes, under physiological conditions. Here, we introduce a novel isobaric crosslinker, Qlinker, for studying conformational and structural changes in proteins and protein complexes using quantitative crosslinking mass spectrometry. Qlinkers are small and simple, amine-reactive molecules with an optimal extended distance of ~10 Å, which use MS2 reporter ions for relative quantification of Qlinker-modified peptides derived from different samples. We synthesized the 2-plex Q2linker and showed that the Q2linker can provide quantitative crosslinking data that pinpoints key conformational and structural changes in biosensors, binary and ternary complexes composed of the general transcription factors TBP, TFIIA, and TFIIB, and RNA polymerase II complexes.