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Select past member of Department Dr Stephen Bell Prof. Jonathan Blackburn Dr David Brown Dr Pak-Lee Chau Dr Hal Dixon (deceased) Dr Jessica Downs Prof. David Ellar Dr Paul Kemp Dr Simon Lovell Dr Kenji Mizuguchi Dr Massimo Paoli Prof. Richard Perham Dr Peter Reynolds Dr Philip Rubery Dr Gerry Smith Dr Margaret Sunde Prof. Jamie Vandenberg Dr Kira Weissman

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Protein complexes in control of gene expression

Research Grouping: Structural and molecular cell biology

My interests lie in understanding the protein structure-function relationships of the systems that control gene expression via chromatin structure.

Figure 1.  Family of the chromo related domains. Equivalent strands are shown using the same colour scheme. Spheres indicate the positions of the conserved aromatic residues involved in methyl-lysine binding. The PDB accession numbers for the structures used in the Figure are as follows: HP1 chromo domain/histone H3 complex – 1guw; HP1 shadow chromo domain/CAF-1 p150 complex – 1s4z; MOF chromo barrel domain – 2bud; MBT domain – 1oi1; Tudor domain – 1mhn; Sso7/DNA complex – 1bf4.

Together with my colleagues in Prof. Ernest Laue’s group I characterized the structures and the interactions of several chromo domain proteins. One of these, heterochromatin protein 1 (HP1), is involved in gene silencing by heterochromatin. Our work led to a functional model, in which the N-terminal chromo domain is required for stable binding of HP1 proteins to methylated histone H3, while other complexes are recruited by binding to the C-terminal shadow chromo domain to act at this specific location in chromatin (1,2).

Based on the structures of several domains we analysed evolutionary relationships within the extended chromo-related domain family.  The structure of the MOF "chromo barrel" domain represents a “missing” link in the superfamily of the chromatin-associated modules, which include the chromo, MBT repeat, Tudor and PWWP domains.  The similarity of the structure, with its clear sequence and structural conservation with the chromo domain, to the common fold of other chromatin-associated modules supports the hypothesis that all these domains share a common ancestry (Figure 1 and (3)).

We identified one of the proteins interacting with the chromo shadow domain of HP1 as the large subunit of chromatin assembly factor 1 (CAF1) (Figure2 and (1)). The CAF1 complex consists of three subunits (p150, p60 and p48) and it assembles nucleosomes into newly synthesized DNA during both replication and nucleotide excision repair. CAF1 is the only known histone chaperone that works in connection with DNA replication. This work shows an important new link between replication and the formation of repressive chromatin structures. Our ultimate aim is to understand how CAF-1 works to assemble histones into nucleosomes. We expect that extending our studies of CAF-1 with both HP1 and histones will illuminate its function. 

This project is a close collaboration with Ernest Laue and Ben Luisi (this Department), Alexey Murzin (MRC Centre for Protein Engineering, Cambridge, UK) and Alain Verreault (IRIC, Universite de Montreal, Canada).

Figure 2.  Identification of the HP1 interacting proteins.  A: CAF1p150 binds to the chromo shadow domain of HP1. A number of overlapping cDNA clones encoding portions of mouse CAF1p150 were isolated through a two-hybrid screen using HP1 as bait. These clones had a 66-residue sequence in common that was expressed and used in limited proteolysis experiments with HP1 chromo shadow domain to define the minimum interacting fragment. The CAF1p150 residues coloured red are involved in hydrophobic interactions with the shadow domain.  B: Proteins identified in the two-hybrid screen contain the PXVXL motif.

Lab members References Thiru A., Nietlispach D., Mott H.R., Okuwaki M., Lyon D., Nielsen P., Hirshberg M., Verreault A., Murzina N.V,* Laue E.D.* (2004) Structural basis of HP1/PXVXL motif peptide interactions and HP1 localisation to heterochromatin. EMBO J., v. 23, 489-99. Epub 2004 Feb 05. (*joint correspondent authors) Nielsen P. R., Nietlispach  D., Mott H. R., Callaghan J.,  Bannister A., Kouzarides T., Murzin A.G,   Murzina N.V.* and Laue E.D.* (2002) Structure of the HP1 chromodomain bound to lysine 9-methylated histone H3. Nature, v.416, 103-7, DOI 10.1038/nature722 (*joint correspondent authors) Nielsen P.R., Nietlispach D., Buscaino A., Warner R.J., Akhtar A., Murzin A.G., Murzina N.V.*, Laue E.D.* (2005) Structure of the chromo barrel domain from the MOF acetyltransferase. J. Biol. Chem., v. 280, 32326-31. Epub 2005 Jun 17.  (*joint correspondent authors)