Investigating the role of Histone Deacetylase 1 in embryonic stem cells and early embryonic development

Histone deacetylase 1 (HDAC1) functions as the catalytic core of multiprotein corepressor complexes removing acetyl marks on histone proteins, causing the compaction of chromatin. Despite this repressive function, the removal of HDAC1 and its highly homologous sister protein HDAC2 leads to the downregulation of large numbers of genes in embryonic stem cells (ESCs) and other cell types. Early embryonic development is one of the few examples where HDAC1 and 2 are not redundant as Hdac1 knockout (KO) mice embryos fail to develop past embryonic day (e) 10.5, whereas Hdac2 KO embryos survive at least until birth. The work in this thesis was conducted with the aim of further unravelling the role of HDAC1 in ESCs and early embryonic development. The proximity-dependent biotinylation (PDB) techniques, APEX2 and BioID, were used to investigate the local protein environment of HDAC1 in ESCs, also facilitating a direct comparison of the two techniques. PDB revealed that the ‘neighbours’ of HDAC1 are involved in many nuclear processes including DNA methylation, while there was a strong overlap in the HDAC1 proximal proteins identified by BioID and APEX2. Epiblast like stem cell (EpiLSC) differentiation was used to study what effect reduced HDAC1/2 levels have on an early stage of embryonic development (roughly e3.5-6.5), revealing the dysregulation of hundreds of genes even at this early time point. The degradation tag (dTag) system was used to allow for far more rapid HDAC1 protein degradation than is seen following conditional KO of Hdac1, revealing that the acute degradation of HDAC1 causes an increase in histone acetylation on specific sites within two hours. RNA-seq showed that there is largely an upregulation of gene expression seen 2 hours following HDAC1 degradation with downregulation becoming more prominent by 24 hours.