Determining cellular CTCF and cohesin abundances to constrain 3D genome models

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  • Claudia Cattoglio
    Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, CIRM Center of Excellence, University of California, Berkeley, Berkeley, United States
  • Iryna Pustova
    Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, CIRM Center of Excellence, University of California, Berkeley, Berkeley, United States
  • Nike Walther
    Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany
  • Jaclyn J Ho
    Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, CIRM Center of Excellence, University of California, Berkeley, Berkeley, United States
  • Merle Hantsche-Grininger
    Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany
  • Carla J Inouye
    Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, CIRM Center of Excellence, University of California, Berkeley, Berkeley, United States
  • M Julius Hossain
    Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany
  • Gina M Dailey
    Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, CIRM Center of Excellence, University of California, Berkeley, Berkeley, United States
  • Jan Ellenberg
    Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany
  • Xavier Darzacq
    Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, CIRM Center of Excellence, University of California, Berkeley, Berkeley, United States
  • Robert Tjian
    Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, CIRM Center of Excellence, University of California, Berkeley, Berkeley, United States
  • Anders S Hansen
    Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, CIRM Center of Excellence, University of California, Berkeley, Berkeley, United States

Description

<jats:p>Achieving a quantitative and predictive understanding of 3D genome architecture remains a major challenge, as it requires quantitative measurements of the key proteins involved. Here, we report the quantification of CTCF and cohesin, two causal regulators of topologically associating domains (TADs) in mammalian cells. Extending our previous imaging studies (Hansen et al., 2017), we estimate bounds on the density of putatively DNA loop-extruding cohesin complexes and CTCF binding site occupancy. Furthermore, co-immunoprecipitation studies of an endogenously tagged subunit (Rad21) suggest the presence of cohesin dimers and/or oligomers. Finally, based on our cell lines with accurately measured protein abundances, we report a method to conveniently determine the number of molecules of any Halo-tagged protein in the cell. We anticipate that our results and the established tool for measuring cellular protein abundances will advance a more quantitative understanding of 3D genome organization, and facilitate protein quantification, key to comprehend diverse biological processes.</jats:p>

Journal

  • eLife

    eLife 8 2019-06-17

    eLife Sciences Publications, Ltd

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