Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA binding
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-wid...
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2025-02-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-025-56515-4 |
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| author | 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. Gebhardt |
| author_facet | 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. Gebhardt |
| author_sort | Duyen Huynh |
| collection | DOAJ |
| description | 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. |
| format | Article |
| id | doaj-art-894d715da444404996be6848429d6bdc |
| institution | Kabale University |
| issn | 2041-1723 |
| language | English |
| publishDate | 2025-02-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj-art-894d715da444404996be6848429d6bdc2025-02-02T12:32:18ZengNature PortfolioNature Communications2041-17232025-02-0116111510.1038/s41467-025-56515-4Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA bindingDuyen Huynh0Philipp Hoffmeister1Tobias Friedrich2Kefan Zhang3Marek Bartkuhn4Francesca Ferrante5Benedetto Daniele Giaimo6Rhett A. Kovall7Tilman Borggrefe8Franz Oswald9J. Christof M. Gebhardt10Institute of Experimental Physics and IQST, Ulm UniversityClinic of Internal Medicine I, University Medical Center UlmInstitute of Biochemistry, Justus-Liebig-Universität GießenInstitute of Experimental Physics and IQST, Ulm UniversityBiomedical Informatics and Systems Medicine, Justus-Liebig-Universität GießenInstitute of Biochemistry, Justus-Liebig-Universität GießenInstitute of Biochemistry, Justus-Liebig-Universität GießenDepartment of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati College of MedicineInstitute of Biochemistry, Justus-Liebig-Universität GießenClinic of Internal Medicine I, University Medical Center UlmInstitute of Experimental Physics and IQST, Ulm UniversityAbstract 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.https://doi.org/10.1038/s41467-025-56515-4 |
| spellingShingle | 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. Gebhardt Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA binding Nature Communications |
| title | Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA binding |
| title_full | Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA binding |
| title_fullStr | Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA binding |
| title_full_unstemmed | Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA binding |
| title_short | Effective in vivo binding energy landscape illustrates kinetic stability of RBPJ-DNA binding |
| title_sort | effective in vivo binding energy landscape illustrates kinetic stability of rbpj dna binding |
| url | https://doi.org/10.1038/s41467-025-56515-4 |
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