1.) Exploring the principle of chromatin sculpting by ubiquitylation.
The recent finding that the spatial chromatin distribution of an HP1-bound anti-silencer factor is controlled through ubiquitin-dependent degradation gave rise to a novel concept of chromatin regulation coined chromatin sculpting (Braun et al., 2011). Here, a boundary factor with anti-silencing activity, Epe1, is uniformly recruited to heterochromatin via binding to HP1 proteins but then selectively removed through ubiquitin-dependent degradation by the ubiquitin ligase complex Cul4-Ddb1Cdt2. Degradation takes place specifically in the body of heterochromatin, which results in the accumulation of Epe1 at the transitions between euchromatin and heterochromatin ('sculpting'; figure A). This spatial regulation prevents the invasion of Epe1 into heterochromatin and specifies its function as a boundary factor.
Intriguingly, other ubiquitin ligases have been identified that are critical for heterochromatin formation. This raises the question whether they follow the same principle of chromatin sculpting and shape the distribution of chromatin-associated factors also through protein degradation (figure B). So far, the specific functions of these ubiquitin ligases in heterochromatin formation remain elusive since the substrates have not been identified. Our goal is to identify these substrates using a combined strategy of proteomic and genomic approaches and to understand how these regulatory pathways contribute to controlling the heterochromatic platform and shaping of chromatin domains.
2.) Dissecting the regulatory networks of heterochromatin formation
Besides ubiquitylation, other molecular mechanisms contribute to the regulatory networks that control the heterochromatic platform. For example, phosphorylation of HP1 proteins (like Swi6 in fission yeast) controls the spatial distribution of HP1-associated factors and various steps in heterochromatin formation such as H3K9 methylation are temporally controlled during the cell cycle. However the underlying molecular mechanisms that specify this spatial and temporal control still need to be identified. Using genetic screens and functional genetics (E-MAP), our goal is to identify novel epigenetic regulators and to dissect the regulatory pathways and networks that contribute to this spatiotemporal control. In addition, we aim to understand the role of plasticity and dynamics in heterochromatin formation by developing new techniques that allow us monitoring silencing at the single cell level.