Chromatin as an expansive canvas for chemical biology

Abstract

Chromatin is extensively chemically modified and thereby acts as a dynamic signaling platform controlling gene function. Chromatin regulation is integral to cell differentiation, lineage commitment and organism development, whereas chromatin dysregulation can lead to age-related and neurodegenerative disorders as well as cancer. Investigating chromatin biology presents a unique challenge, as the issue spans many disciplines, including cell and systems biology, biochemistry and molecular biophysics. In recent years, the application of chemical biology methods for investigating chromatin processes has gained considerable traction. Indeed, chemical biologists now have at their disposal powerful chemical tools that allow chromatin biology to be scrutinized at the level of the cell all the way down to the single chromatin fiber. Here we present recent examples of how this rapidly expanding palette of chemical tools is being used to paint a detailed picture of chromatin function in organism development and disease.

Figure 1. Organization and analysis of chromatin

(a) Schematic overview of the structural organization of chromatin highlighting the type of chemical biology tools that have been successfully used to explore each level of complexity. For the structure of the nucleosome, see Protein Data Bank (PDB) code 1kx3. MN, mononucleosome. (b) Summary of the major chromatin states with their associated histone marks indicated. me, methylation; ub, ubiquitylation; ac, acetylation.

Figure 2. Chemical synthesis of designer chromatin

(a–c) PTMs and their analogs introduced into histone proteins by EPl or NCl methods (a), modification of single cysteine residues (b) and nonsense suppression methods (c, for details see text). Kme1, monomethylated lysine; Kme2, dimethylated lysine; Kme3, trimethylated lysine; Kac, acetylated lysine; Kub, lysine ubiquitylation; o-ADPR, o-ADP ribosylation; Sph, phosphorylated serine; pHis, phosphorylated histidine; pTza, phosphoryltriazolylalanine. (d) Methods to reconstitute designer chromatin of different lengths and compositions. NPS, nucleosome positioning sequence; Nap-1, nucleosome assembly protein 1.

Figure 3. Small-molecule inhibitors of writers and erasers of histone marks

(a) Structures of HDAC inhibitors with their known targets indicated. (b) Structures of HMT inhibitors and their targets.

Figure 4. Readout of histone marks

(a) Structures of representative acetyllysine and methyllysine reader domains. Each domain is shown in complex with a histone-derived modified peptide. Shown are the GCN5 bromodomain (PDB code 1e6i), the PHD finger from BPTF (PDB code 2f6j), the HP1 chromodomain (PDB code 1kne), the MBT domain of l3MBTl1 (PDB code 2pqw) and the HTD of JMJD2A (PDB code 2gfa). (b) Schematic representation of the various ways reader domains can be used to readout chromatin states. As discussed in the text, many domains can bind to multiple sites on histones (each carrying the same modification, left). Specificity can be imposed through the presence of nearby heterotypic marks that impede binding (middle) or through multivalent interactions involving homotypic or heterotypic marks on either the same histone or different histones in a nucleosome (right).

Figure 5. Small-molecule inhibitors of histone-binding modules

Chemical structures of the small molecules that target the peptide-binding pockets of various histone reader domains. See the text for details. BET-bromo, BET bromodomain; PCAF-bromo, PCAF bromodomain; CBP-bromo, CBP bromodomain; l3MBTl1-MBT, l3MBTl1 MBT domain.

Figure 6. Control of chromatin function by histone marks through cis-and trans-acting mechanisms

The combination of designer chromatin and various biochemical and biophysical methods allows for a deeper understanding of the ways in which histone marks can locally regulate chromatin structure and function. (a–c) Examples of histone mark crosstalk. Such crosstalk can occur when a chromatin effector is recruited to chromatin by a specific mark and then proceeds to install additional copies of the same mark, as in the cooperative acetylation of chromatin by the SAGA complex. Alternatively, an effector is recruited by a mark that is distinct from the one it then acts on, as observed in the Rdp3S HDAC complex. Allosteric crosstalk between marks was determined for the H3K79 methyltransferase Dot1, which is activated by H2B ubiquitylation. (d) Certain histone marks intrinsically alter the structural properties of chromatin fibers: H4K20me3 leads to the stabilization of a compact fiber structure, whereas H4K16ac and uH2B impair fiber compaction. (e) Histone marks can alter the stability of a single nucleosome: specific acetylation sites in the globular domain of H3 and H4 modulate nucleosomal stability and lead to increased DNA access to downstream effectors.