Characterising the role and mechanism of action of the MIER1:HDAC complex in gene repression and X chromosome inactivation

The appropriate regulation of our genes is essential for controlling embryonic development as well as enabling adult organisms to respond to both internal and external challenges including metabolism and disease. Our genes are regulated by many large protein complexes that often control post-translational chromatin modifications. In this way these complexes can control chromatin organization which in turn controls gene accessibility and activity. One important family of complexes contain enzymes that remove acetyl groups from the chromatin. These so-called “class I histone deacetylase enzymes”, HDACs 1,2&3 form the catalytic subunits of seven distinct genome-targeted complexes. The importance of these HDAC complexes is highlighted by the fact that there are currently more than 100 active clinical trials of HDAC inhibitors (clinicaltrials.gov) and five inhibitors of class I HDACs have been licensed for the treatment of cutaneous T cell lymphoma, multiple myeloma and epilepsy. Four of these seven distinct HDAC complexes (NuRD, CoREST, SIN3A/B and SMRT/NCoR) have well-established roles in controlling gene expression and are recruited to many loci across the genome. A further three complexes, the MIER, MiDAC and RERE complexes, are less well understood. In this application we propose to study the MIER histone deacetylase complex, which is clearly important since it is conserved from worms to man. We have found that removing the MIER complex in a cancer cell line has no effect on gene expression. This is quite distinct from other HDAC complexes and suggests that the complex may play a role in other processes – perhaps during development. Fitting with this, we have observed, in HEK293 cells, that the MIER1 complex is heavily enriched on the inactive X-chromosome. The inactivation of one X-chromosome is important in female cells to ensure the correct “gene dosage” compared with male cells that have only a single X-chromosome. It is therefore important that we understand what role the MIER complex plays in establishing heterochromatin formation that results in gene silencing. We have begun biochemical studies of the MIER complex which show that it associates with nucleosomes bearing the repressive histone mark H3K27me2/3 through the BAHD1 component. We expect that it will also recognise H3K9me2 through the CDYL component. We have also shown that it binds tightly to a H2A:H2B histone dimer. We will address two key research questions to understand the MIER complex: (i) How is the MIER:HDAC complex assembled and how does it target its chromatin substrate? What is the structure of the MIER1:HDAC1 complex and how does it compare with other HDAC complexes? How does the MIER1:HDAC1 complex interact with chromatin and how do the different components contribute to the specificity of interaction? (ii) What is the role of the complex in X-chromosome inactivation? When and how is MIER:HDAC recruited to the X-chromosome? What does the complex do to contribute to X-chromosome inactivation? To address these questions we will apply our collective expertise in biochemistry, structural biology, quantitative super-resolution microscopy and gene editing. We utilise our state of the art cryoEM facility to deliver the structural biology. We have recently installed a new BBSRC-funded super-resolution microscope to understand the cell biology in both time and space. Overall this project will provide a vastly improved understanding of the MIER complex in terms of both structure and mechanism of action and the biological role of the MIER complex in X-chromosome inactivation.