FACT weakens the nucleosomal barrier to transcription and preserves its integrity by forming a hexasome-like intermediate

Francesca Burgos-Bravo¹, Alexander B Tong², Chen Li³, César Díaz-Celis⁴, Craig D Kaplan⁵, Gary LeRoy⁶, Danny Reinberg⁶, Carlos Bustamante⁷

Affiliations

¹ Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
² Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Chemistry, University of California, Berkeley, Berkeley, CA 94720, USA.
³ Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Biophysics Graduate Group, University of California, Berkeley, Berkeley, CA 94720, USA.
⁴ Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
⁵ Department of Biological Sciences, University of Pittsburgh, Pittsburgh, CA 15260, USA.
⁶ Howard Hughes Medical Institute, University of Miami Miller School of Medicine, Miami, FL 33136, USA.
⁷ Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA; Biophysics Graduate Group, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Chemistry, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Physics, University of California, Berkeley, Berkeley, CA 94720, USA; Molecular Biophysics and Integrative Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA; Kavli Energy Nanoscience Institute, University of California Berkeley, Berkeley, CA 94720, USA. Electronic address: carlosb@berkeley.edu.

PMID: 40412388
PMCID: PMC12204072 (available on 2026-06-05)
DOI: 10.1016/j.molcel.2025.05.002

FACT weakens the nucleosomal barrier to transcription and preserves its integrity by forming a hexasome-like intermediate

Francesca Burgos-Bravo et al. Mol Cell. 2025.

. 2025 Jun 5;85(11):2097-2109.e8.

doi: 10.1016/j.molcel.2025.05.002. Epub 2025 May 23.

Authors

Francesca Burgos-Bravo¹, Alexander B Tong², Chen Li³, César Díaz-Celis⁴, Craig D Kaplan⁵, Gary LeRoy⁶, Danny Reinberg⁶, Carlos Bustamante⁷

Affiliations

¹ Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
² Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Chemistry, University of California, Berkeley, Berkeley, CA 94720, USA.
³ Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Biophysics Graduate Group, University of California, Berkeley, Berkeley, CA 94720, USA.
⁴ Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
⁵ Department of Biological Sciences, University of Pittsburgh, Pittsburgh, CA 15260, USA.
⁶ Howard Hughes Medical Institute, University of Miami Miller School of Medicine, Miami, FL 33136, USA.
⁷ Jason Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, CA 94704, USA; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94704, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA; Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA; Biophysics Graduate Group, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Chemistry, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Physics, University of California, Berkeley, Berkeley, CA 94720, USA; Molecular Biophysics and Integrative Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA; Kavli Energy Nanoscience Institute, University of California Berkeley, Berkeley, CA 94720, USA. Electronic address: carlosb@berkeley.edu.

PMID: 40412388
PMCID: PMC12204072 (available on 2026-06-05)
DOI: 10.1016/j.molcel.2025.05.002

Abstract

Transcription of yeast RNA polymerase II through nucleosomes requires the assistance of the histone chaperone FACT (facilitates chromatin transcription). Yet, how FACT modulates the nucleosomal mechanical barrier to affect the polymerase's elongation dynamics is poorly understood. Using high-resolution single-molecule optical tweezers, we show that FACT greatly decreases the magnitude of the barrier by favoring the unwrapping of DNA from the distal H2A-H2B dimer, which, in turn, weakens the contacts near the dyad, significantly reducing the enzyme's crossing time. We show that barrier crossing depends on the asymmetric flexibility of the nucleosome arms, an asymmetry we find across the genome. Mechanical unwrapping of Cy3-H2A nucleosomes reveals that FACT reduces their unwrapping force and stabilizes a hexasome-like intermediate that retains both labeled dimers during successive unwrapping cycles. This intermediate is also observed after transcription. In conclusion, FACT facilitates nucleosomal transcription by weakening the barrier and actively assisting the maintenance of nucleosomal integrity after enzyme passage.

Keywords: FACT; RNA polymerase II; histone chaperone; nucleosomal DNA flexibility; nucleosome; nucleosome integrity; nucleosome unwrapping; optical tweezers; single-molecule fluorescence; transcription.

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Conflict of interest statement

Declaration of interests The authors declare no competing interests.

References

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FACT weakens the nucleosomal barrier to transcription and preserves its integrity by forming a hexasome-like intermediate

Affiliations

FACT weakens the nucleosomal barrier to transcription and preserves its integrity by forming a hexasome-like intermediate

Authors

Affiliations

Abstract

Conflict of interest statement

References

MeSH terms

Substances

Grants and funding

LinkOut - more resources

Full Text Sources