Achievements

Our molecular structures of the nucleosome core, the tetranucleosome, a chromatin remodeling factor and transcription factors lay the foundation for the mechanistic understanding of DNA-dependent, eukaryotic nuclear processes. Please see the descriptions below and selected references.

Nucleosome Core

The nucleosome is the fundamental repeating unit of chromatin, and both its internal and higher-order structures are crucial to the functioning of DNA in the nucleus. We solved the X-ray structure of the nucleosome core originally at 2.8 Å resolution (Figure: Nucleosome Core) [Luger et al.] and ultimately at 1.9 Å resolution (Figure: Nucleosome Core 1.9A) [Davey et al.]. This structure contains 147 base pairs of DNA and two copies of each of the four core histone proteins. This highly reliable structure shows, for example, all the direct and water-mediated hydrogen bonds between protein and DNA molecules. A detailed analysis of the DNA reveals sequence specific conformations [Richmond and Davey].

Luger K., A. W. Mäder, R. K. Richmond, D. F. Sargent, and T. J. Richmond (1997) "Crystal structure of the nucleosome core particle at 2.8 Å resolution", Nature 389, 251.

Davey C. A., D. F. Sargent, K. Luger, A. W. Mäder, and T. J. Richmond (2002) "Solvent mediated interactions in the structure of the nucleosome core particle at 1.9 Å resolution", J Mol Biol 319, 1097.

Richmond T. J., and C. A. Davey (2003) "The structure of DNA in the nucleosome core", Nature 423, 145.

Tetranucleosome

In their most condensed state, arrays of nucleosomes are helically arranged in a fiber approximately 30 nm in diameter. To understand the structural organization of this nucleosome ‘higher-order’ structure, we crystallized a tetranucleosome and determined its X-ray structure at 9 Å resolution [Schalch et al.]. The structure shows two stacks of nucleosomes with linker DNA running between them (Figure: Tetranucleosome 9A), and is compatible with a two-start, but not a one-start helix (i.e. the solenoid). This result corresponds well with our biochemical and electron microscopy analyses also showing a two-start arrangement [Dorigo et al.]. A model of the chromatin fiber built by stacking tetranucleosomes on each other is consistent with the properties of chromatin fragments (Figure: Chromatin Fiber).

Schalch T., S. Duda, D. F. Sargent, and T. J. Richmond (2005) "X-ray structure of a tetranucleosome and its implications for the chromatin fibre", Nature 436, 138.

Dorigo B., T. Schalch, A. Kulangara, S. Duda, R. R. Schroeder, and T. J. Richmond (2004) "Nucleosome Arrays Reveal the Two-Start Organization of the Chromatin Fiber", Science 306, 1571.

ISW1a/dinucleosome Model

Nucleosomes are not typically static entities fixed along genomic DNA, but are capable of repositioning and thereby altering the accessibility of transcription regulatory sites and gene-coding regions. Although mobility is a property intrinsic to nucleosomes, the rate of translocation is evidently too slow to accommodate most processes in the nucleus. ATP-dependent, multiprotein complexes called chromatin remodeling factors can dislodge, reconfigure and translocate nucleosomes.

We solved the X-ray structure of the yeast ISW1a chromatin remodeling factor lacking its ATPase domain, both alone and with a 48 bp DNA bound at two sites (resolutions of 3.2 and 3.6 Å, respectively) [Yamada et al.]. Using cryo-electron microscopy, we visualized this ISW1a construct in two different nucleosome complexes. The composite X-ray and electron microscopy structures combined with site-directed photocrosslinking analyses suggest that ISW1a uses a dinucleosome substrate for chromatin remodeling (Figure: ISW1a/dinucleosome Model]. Results from a remodeling assay corroborate the dinucleosome model. Our study shows how ISW1a could set the spacing between two adjacent nucleosomes by acting as a ‘protein ruler’.

Figure note: The ATPase domain of the Isw1 protein (brown) is taken from Thoma et al., Nat Struc Biol 2005, and placed in the model using the footprinting results of Gangaraju and Bartholomew, Mol Cell Biol 2007.

K. Yamada, T. D. Frouws, B. Angst, D. J. Fitzgerald, C. DeLuca, K. Schimmele, D. F. Sargent and T. J. Richmond (2011). "Structure and mechanism of the chromatin remodelling factor ISW1a", Nature 472, 448-53.

More to come - under construction.