Report

Teaser, summary, work performed and final results

Periodic Reporting for period 2 - PolyDomFormFuncReg (Discovering how polycomb domains form and function in gene regulation)

Teaser

The capacity to control gene expression is fundamental to normal cellular function and organismal development. When these processes go wrong this can lead to debilitating human disease. Through studying the mechanisms by which the cell regulates gene expression it has become...

Summary

The capacity to control gene expression is fundamental to normal cellular function and organismal development. When these processes go wrong this can lead to debilitating human disease. Through studying the mechanisms by which the cell regulates gene expression it has become clear that chromatin, the template on which gene transcription occurs, can profoundly affect gene expression. In studying this aspect of gene expression, it has been shown that post-translation modifications to histones are central to gene regulation by chromatin. The polycomb repressive proteins form a highly conserved chromatin modifying system in animals. The polycomb system is known to repress gene expression, but how polycomb proteins are recruited to target genes and ultimately how they create chromatin environments that are repressive to gene expression remains poorly understood. In the context of understanding polycomb mediated gene repression, the central aims of this ERC funded programme are to (i) Discover how the KDM2B/PRC1 complex initiates polycomb domain formation, (ii) Discover how polycomb target sites are selected and polycomb domains formed during normal cell lineage commitment, and (iii) Discover how polycomb domains regulate gene expression. In progressing these aims, the programme has already made significant advances. We have discovered that the KDM2B/PRC1 protein complex is highly active at placing a modification on chromatin (H2AK119ub1) in vitro and that this activity is regulated by a protein called RYBP (Rose NR, et al, 2016). Together this KDM2B/PRC1 activity and its regulation by RYBP appear to be important for how polycomb chromatin domains form and function inside cells. Building on this discovery, we have now pinpointed the distinct polycomb repressive complexes that are responsible for gene repression and identified the primary target genes that the polycomb system regulates. Interestingly, we discovered that repression by the polycomb system does not appear to control access to DNA as has previously been postulated (King HW, et al, 2018). Together these discoveries are forming the basis on which we will continue to better understand how the polycomb repressive system functions during cell lineage commitment. Building on these discoveries, we will continue to define how the polycomb system functions in cell lineage commitment and potentially provide new avenues for therapeutic intervention in human diseases where the polycomb system is perturbed.

Work performed

Since the beginning of the project we have assembled a talented team of researchers to address its central aims.
We have explored how polycomb chromatin domains are formed (Aim 1) using biochemical approaches and discovered that KDM2B/PRC1 is highly active at placing H2AK119ub1 and that this activity is regulated. This was an important new advance as it demonstrated that KDM2B/PRC1, based on its in vitro activity, may play a unique role in regulating H2AK119ub1 in the cell. Given that we had showed in our previous work, using model systems, that H2AK119ub1 was required for polycomb chromatin domain formation, we then went on to show inside cells that the mechanisms that simulate and regulate H2AK119bu1 through KDM2B/PRC1 are required for normal polycomb chromatin domain formation. This work was published in the journal eLIFE.
In the context of Aim 2, we have been working towards understanding how polycomb chromatin domains form during cell lineage commitment. To achieve this we have initially been systematically dissecting the biochemical features of polycomb chromatin domains in vivo and defining the PRC1 complexes that contribute to polycomb chromatin domain formation in progenitor cells. This is required to narrow down where to focus our efforts examining these processes during cell lineage commitment. This has been an essential step in our progress on this aim given the complexity of PRC1 complexes that has emerged recently from our own work and the literature. In the context of this work, we have revealed that polycomb proteins do not limit accessibility to the underlying DNA as a means of gene repression, thereby disproving a view that has been long proposed in the field. Instead it suggests that polycomb chromatin domains must contribute to gene repression through alternative mechanisms. This important new finding was published in Genome Research. Towards defining the contribution of individual PRC1 complexes to polycomb chromatin domain formation we are in the process of systemically deleting individual complexes and examining how they affect polycomb chromatin domain formation and gene repression. We have made interesting and important new observations in the context of these experiments which are essential to meeting the goals of this aim.
In discovering how polycomb chromatin domains regulate gene expression (Aim 3) we have now identified primary target genes of PRC1 and are in the process of discovering the mechanisms by which polycomb chromatin domains affect gene transcription.
Together these new discoveries are beginning to uncover how polycomb repressive systems regulate gene expression in mammals.

Final results

As we integrate the emerging discoveries from the three central aims of this project we are achieving unprecedented new understanding into the molecular mechanisms that define biochemically how polycomb chromatin domains form in vitro, how this is achieved inside cells during cell lineage commitment, and the molecular mechanisms that ultimately lead to polycomb-mediated repression of gene transcription. Our biochemical dissection of the PRC1 complex is the most complete aspect of our progress to date, so the remainder of the project will concentrate on the latter two aims which are focussed on the in vivo function of the polycomb system. This will provide essential new insight into how the polycomb repressive systems regulate gene expression in mammals.