Regulation of Chromatin by Combinatorial DNA and Histone Modifications

Final Report Summary - ROCCOMO (Regulation of Chromatin by Combinatorial DNA and Histone Modifications)

Most of the genetic material of eukaryotic cells is stored in the nucleus in the form of chromatin, in which the DNA is packaged with the help of histones and other regulatory and structural proteins. DNA and histones carry chemical modifications that play central roles in regulating chromatin function as they participate in all DNA-related processes, such as transcription, DNA replication, and DNA repair. These modifications are deposited by epigenetic ‘writers’ and recognised by epigenetic ‘readers’ that bind to these modifications through specialised binding domains. Mutations in epigenetic readers and writers and the resulting errors in chromatin regulation often result in diseases such as neuronal disorders or cancer.
It is thought that DNA and histone modifications form an ‘epigenetic code’ on chromatin that stores epigenetic information in addition to the genetic information contained in the DNA sequence. The ‘epigenetic code’ theory, however, has not been systematically tested, and it was not clear whether chromatin modifications indeed form a code and how this code could operate. The goal of the ROCCOMO project was to investigate how the epigenetic code works and to find out how epigenetic regulators extract information from chromatin by ‘reading out’ combinations of DNA and histone modifications.
Towards this goal we have developed chemical biology methods to make artificial ‘designer’ chromatin containing the desired DNA and histone modifications. We have created a library of different chromatin types that resemble active ‘promoter’ or ‘enhancer’ or repressed ‘heterochromatin’ states. Using this library of chromatin states as ‘bait’ in affinity purifications we have characterised the binding of ~1500 nuclear proteins to the various forms of modified chromatin via quantitative mass spectrometry measurements. New bioinformatic tools that we have developed to analyse the mass spectrometry data has allowed us to define different binding modes of these proteins to the modified chromatin. While many proteins have binding characteristics that can be explained by simple rules defined by binding or exclusion induced by one or a few modifications, other proteins show complex binding modes that cannot be explained by simple modification ‘codes’. Our investigation of selected epigenetic regulators further revealed that features other than modifications, such chromatin conformation (Foster et al., 2018, Mol. Cell 72:739) or accessibility and additional proteins (Borgel et al., 2017, Nucleic Acids Res. 45:1114) might be cues that are recognised by epigenetic readers. We also found that modifications not only regulate chromatin binding but also act by modulating ‘writer’ activities, as we could demonstrate in our analysis of the enzymatic mechanism of the ubiquitin ligase UHRF1 (Foster et al., 2018, Mol. Cell 72:739), a factor that is essential for the maintenance of DNA methylation. Lastly, our experiments have identified the BRCA1 DNA repair complex as an epigenetic reader that recognises newly replicated chromatin. BRCA1 binds to chromatin through the ankyrin repeat domain (ARD) of its BARD1 subunit (Nakamura et al., 2019, Nat. Cell Biol. 21:311). This has important consequences for the development of novel epigenetic inhibitors. Compounds that block the BARD1 ARD from binding chromatin would open up the possibility to chemically inactivate BRCA1, which could be exploited for cancer treatments.
In summary, we have succeeded in describing how chromatin regulators read out different chromatin states, and by investigating how specific epigenetic readers interpret combinatorial DNA and histone modification signatures we could demonstrate principle modes of operation of the epigenetic code. Some of our findings have important implications for understanding how epigenetic processes regulate the integrity of the genome, and devise a path for the development of a novel class of epigenetic drugs for cancer therapies.

Final Report Summary - ROCCOMO (Regulation of Chromatin by Combinatorial DNA and Histone Modifications)

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