Nucleosome Opening Kinetics and the Influence of Histone Modifications Studied by Single Molecule FRET

Nucleosomes compact the genome and regulate access by wrapping and unwrapping DNA. Here we characterized NaCl-induced opening of mononucleosomes by single-molecule FRET, using donor/acceptor labels on the DNA and various histones. By species-selective fluorescence lifetime and photon distribution analysis we identified new nucleosome opening intermediates and developed a kinetic model [1]. Opening proceeds through a weakening of the H2A-H2B dimer/(H3-H4)2 tetramer interface on a 0.1 ms time scale, then by a slower two-step release of the dimers coupled to DNA unwrapping, extending from several ms to minutes. Nucleosome opening and detachment of histone dimers proceed asymmetrically depending on the DNA sequence.Mutations at the H2A/H3 interface (H2A R81A, R88A, R81A/R88A, R81E/R88E) facilitate the initial opening, confirming the importance of the dimer:tetramer interface for nucleosome stability. This is also supported by molecular dynamics (MD) simulations that show enhanced DNA fluctuations for the mutants. Partially opened states such as described here might be a convenient nucleation point for DNA-recognizing proteins.For characterizing the role of histone tails in nucleosome interaction and stability, we performed MD simulations in explicit and implicit solvent on the H4 tail interacting with the acidic patch of a neighboring nucleosome. The simulations show that H4K16 acetylation decreases the interactions between H4 tail and the acidic patch of the neighboring nucleosome, a process that is central to the regulation of chromatin compaction [2,3].References[1] A. Gansen, A. Valeri, F. Hauger, S. Felekyan, S. Kalinin, K. Toth, J. Langowski, and C. A. M. Seidel. Nucleosome disassembly intermediates characterized by single-molecule FRET. Proc Natl Acad Sci U S A, 106(36):15308-13, Sep 2009.[2] J. Erler, R. Zhang, L. Petridis, X. Cheng, J. C. Smith, and J. Langowski. The role of histone tails in the nucleosome: A computational study. Biophys. J., 107(12):2911-2922, 2014.[3] R. Zhang, J. Erler, J. Langowski. How does histone acetylation regulate chromatin accessibility? A computational study of the role of H4K16 in inter-nucleosome interaction. Biophys. J., under revision.