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Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA binding

Author

Listed:
  • Duyen Huynh

    (Ulm University)

  • Philipp Hoffmeister

    (University Medical Center Ulm)

  • Tobias Friedrich

    (Justus-Liebig-Universität Gießen
    Justus-Liebig-Universität Gießen
    Institute for Lung Health (ILH))

  • Kefan Zhang

    (Ulm University)

  • Marek Bartkuhn

    (Justus-Liebig-Universität Gießen
    Institute for Lung Health (ILH))

  • Francesca Ferrante

    (Justus-Liebig-Universität Gießen)

  • Benedetto Daniele Giaimo

    (Justus-Liebig-Universität Gießen)

  • Rhett A. Kovall

    (University of Cincinnati College of Medicine)

  • Tilman Borggrefe

    (Justus-Liebig-Universität Gießen)

  • Franz Oswald

    (University Medical Center Ulm)

  • J. Christof M. Gebhardt

    (Ulm University)

Abstract

Transcription factors (TFs) such as RBPJ in Notch signaling bind to specific DNA sequences to regulate transcription. How TF-DNA binding kinetics and cofactor interactions modulate gene regulation is mostly unknown. We determine the binding kinetics, transcriptional activity, and genome-wide chromatin occupation of RBPJ and mutant variants by live-cell single-molecule tracking, reporter assays, and ChIP-Seq. Importantly, the search time of RBPJ exceeds its residence time, indicating kinetic rather than thermodynamic binding stability. Impaired RBPJ-DNA binding as in Adams-Oliver-Syndrome affect both target site association and dissociation, while impaired cofactor binding mainly alters association and unspecific binding. Moreover, our data point to the possibility that cofactor binding contributes to target site specificity. Findings for other TFs comparable to RBPJ indicate that kinetic rather than thermodynamic DNA binding stability might prevail in vivo. We propose an effective in vivo binding energy landscape of TF-DNA interactions as instructive visualization of binding kinetics and mutation-induced changes.

Suggested Citation

  • Duyen Huynh & Philipp Hoffmeister & Tobias Friedrich & Kefan Zhang & Marek Bartkuhn & Francesca Ferrante & Benedetto Daniele Giaimo & Rhett A. Kovall & Tilman Borggrefe & Franz Oswald & J. Christof M., 2025. "Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA binding," Nature Communications, Nature, vol. 16(1), pages 1-15, December.
  • Handle: RePEc:nat:natcom:v:16:y:2025:i:1:d:10.1038_s41467-025-56515-4
    DOI: 10.1038/s41467-025-56515-4
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