Quantification of protein abundance and interaction defines a mechanism for operation of the circadian clock

  1. Alex Ashton Koch
  2. James S Bagnall
  3. Nicola J Smyllie
  4. Nicola Begley
  5. Antony D Adamson
  6. Jennifer L Fribourgh
  7. David G Spiller
  8. Qing-Jun Meng
  9. Carrie L Partch
  10. Korbinian Strimmer
  11. Thomas A House
  12. Michael H Hastings
  13. Andrew SI Loudon  Is a corresponding author
  1. University of Manchester, United Kingdom
  2. Medical Research Council, United Kingdom
  3. University of California, Santa Cruz, United States

Abstract

The mammalian circadian clock exerts control of daily gene expression through cycles of DNA binding. Here we develop a quantitative model of how a finite pool of BMAL1 protein can regulate thousands of target sites over daily time scales. We used quantitative imaging to track dynamic changes in endogenous labelled proteins across peripheral tissues and the SCN. We determine the contribution of multiple rhythmic processes coordinating BMAL1 DNA binding, including cycling molecular abundance, binding affinities and repression. We find nuclear BMAL1 concentration determines corresponding CLOCK through heterodimerization and define a DNA residence time of this complex. Repression of CLOCK:BMAL1 is achieved through rhythmic changes to BMAL1:CRY1 association and high affinity interactions between PER2:CRY1 which mediates CLOCK:BMAL1 displacement from DNA. Finally, stochastic modelling reveals a dual role for PER:CRY complexes in which increasing concentrations of PER2:CRY1 promotes removal of BMAL1:CLOCK from genes consequently enhancing ability to move to new target sites.

Data availability

Modelling and analtyical code has been made publicly available via GitHub. The FCS analysis software is at https://github.com/LoudonLab/FcsAnalysisPipeline and the modeling link is https://github.com/LoudonLab/CLOCK-BMAL1-DNA-Binding.Source Data files have been provided for all FCS measurements and FRAP measurements in Figures 1, 2, 3 ,4, and 6.

The following data sets were generated

Article and author information

Author details

  1. Alex Ashton Koch

    Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  2. James S Bagnall

    Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  3. Nicola J Smyllie

    Laboratory of Molecular Biology, Medical Research Council, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  4. Nicola Begley

    Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  5. Antony D Adamson

    Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  6. Jennifer L Fribourgh

    Department of Chemistry and Biochemistry, University of California, Santa Cruz, Santa Cruz, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. David G Spiller

    Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  8. Qing-Jun Meng

    Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9426-8336
  9. Carrie L Partch

    Department of Chemistry and Biochemistry, University of California, Santa Cruz, Santa Cruz, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4677-2861
  10. Korbinian Strimmer

    Department of Mathematics, University of Manchester, Manchester, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  11. Thomas A House

    Department of Mathematics, University of Manchester, Manchester, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  12. Michael H Hastings

    Laboratory of Molecular Biology, Medical Research Council, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  13. Andrew SI Loudon

    Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom
    For correspondence
    andrew.loudon@manchester.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3648-445X

Funding

Biotechnology and Biological Sciences Research Council (BB/P017347/1)

  • James S Bagnall
  • Nicola Begley
  • Andrew SI Loudon

Biotechnology and Biological Sciences Research Council (BB/P017355/1)

  • Nicola J Smyllie
  • Michael H Hastings

Medical Research Council (MC_U105170643)

  • Michael H Hastings

National Institutes of Health (GM107069)

  • Carrie L Partch

National Institutes of Health (GM141849)

  • Carrie L Partch

Wellcome Trust (107851/Z/15/Z)

  • Andrew SI Loudon

Wellcome Trust (216416/Z/19/Z)

  • Alex Ashton Koch

University of California

  • Jennifer L Fribourgh

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

Ethics

Animal experimentation: All experimental procedures were carried out in accordance with the Animals (Scientific Procedures) Act of 1986, UK (License number PP7901495).

Copyright

© 2022, Koch 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.

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  1. Alex Ashton Koch
  2. James S Bagnall
  3. Nicola J Smyllie
  4. Nicola Begley
  5. Antony D Adamson
  6. Jennifer L Fribourgh
  7. David G Spiller
  8. Qing-Jun Meng
  9. Carrie L Partch
  10. Korbinian Strimmer
  11. Thomas A House
  12. Michael H Hastings
  13. Andrew SI Loudon
(2022)
Quantification of protein abundance and interaction defines a mechanism for operation of the circadian clock
eLife 11:e73976.
https://doi.org/10.7554/eLife.73976

Share this article

https://doi.org/10.7554/eLife.73976

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