From crystal and NMR structures, footprints and cryo-electron-micrographs to large and soft structures: nanoscale modeling of the nucleosomal stem

Abstract

The interaction of histone H1 with linker DNA results in the formation of the nucleosomal stem structure, with considerable influence on chromatin organization. In a recent paper [Syed,S.H., Goutte-Gattat,D., Becker,N., Meyer,S., Shukla,M.S., Hayes,J.J., Everaers,R., Angelov,D., Bednar,J. and Dimitrov,S. (2010) Single-base resolution mapping of H1-nucleosome interactions and 3D organization of the nucleosome. Proc. Natl Acad. Sci. USA, 107, 9620–9625], we published results of biochemical footprinting and cryo-electron-micrographs of reconstituted mono-, di- and tri-nucleosomes, for H1 variants with different lengths of the cationic C-terminus. Here, we present a detailed account of the analysis of the experimental data and we include thermal fluctuations into our nano-scale model of the stem structure. By combining (i) crystal and NMR structures of the nucleosome core particle and H1, (ii) the known nano-scale structure and elasticity of DNA, (iii) footprinting information on the location of protected sites on the DNA backbone and (iv) cryo-electron micrographs of reconstituted tri-nucleosomes, we arrive at a description of a polymorphic, hierarchically organized stem with a typical length of 20 ± 2 base pairs. A comparison to linker conformations inferred for poly-601 fibers with different linker lengths suggests, that intra-stem interactions stabilize and facilitate the formation of dense chromatin fibers.

Toward Convergence of Experimental Studies and Theoretical Modeling of the Chromatin Fiber

“…Mapping with micrococcal nuclease (29) or hydroxyl radicals (36) provided strong evidence for positioning of LH at the dyad axis with symmetric protection of ϳ11 bp of each linker DNA. This symmetry and the ability of the globular domain alone to bring two linker DNA segments in close juxtaposition to form linker DNA stem motifs are consistent with energy-minimized models (37,38).…”

“…On the basis of a striking homology between the C-terminal domain of LH and the HMG box fold motif, Bharath et al (39) predicted that the folded C-terminal domain could induce formation of linker DNA stems by kinking inward and then diverging at the C-terminal domain. More recent analysis of experimental data and modeling studies have provided a nanoscale model of the LH-induced stem structure (38). Condensation consistent with folding of the C-terminal domain upon H1 binding to the nucleosome was recently con-firmed by FRET experiments (40).…”

Structural insights into the histone H1-nucleosome complex

“…These models are divided into two major classes, symmetric and asymmetric, on the basis of the location of gH1/gH5 in the nucleosome. In the symmetric class, gH1/gH5 binds to the nucleosomal DNA at the dyad and interacts with both linker DNAs (16,17,27,28). In the asymmetric class, gH1/gH5 binds to the nucleosomal DNA in the vicinity of the dyad axis and to 10 bp (27,(29)(30)(31)(32) or 20 bp (19,29,33,34) of one linker DNA, or is located inside the DNA gyres, where it interacts with histone H2A (35).…”

The chromatin fiber: multiscale problems and approaches

“…Although all-atom trajectories of the nucleosome-linker histone complex are too costly, the Wade lab has combined docking, Brownian dynamics, and normal mode analysis to simulate the binding to the H5 globular domain in the nucleosome (without core histone tails) [39 • ]. Their studies suggest that H5 can adopt various docking positions near the nucleosome's dyad axis rather than a single symmetric position where it interacts with both linker DNAs as previously observed [40]. The positional diversity of H5 binding in the nucleosome is important, as it modulates the entering/exiting angles of linker DNAs, which in turn determine higher-order structure of chromatin fibers [41 • ].…”