Investigating the role of Sin3A in embryonic stem cells, during early development and in HCT-116 cells

Class I HDACs are the functional core of several distinct complexes that remove the acetyl marks from histone proteins. We sought to investigate one of the class HDAC complexes, the Sin3 complex, to investigate its role in ES cells and during early development. The core of the Sin3 complex is one of two highly related proteins, either Sin3A or Sin3B. Sin3A has been implicated as the more important protein, as during early embryogenesis as Sin3A-KO (Knockout) mice embryos are not found past embryonic day (e) 6.5, while Sin3B-KO mice embryos show lethality at e14.5. The aim of this thesis is to further investigate the role of the SIN3A complex in ES cells and during early differentiation. We utilised advancements in gene silencing to treat cells with proteolysis targeting chimeras (PROTACs) which resulted in rapid degradation of the SIN3A protein significantly quicker than in previous studies. We show that the loss of SIN3A causes over a thousand genes to become dysregulated within 6 hours and that this is mirrored by changes in the acetylation state of several histone residues. We utilised the gastruloid system to investigate early embryogenesis and we see that gastruloids lacking Sin3A are unable to undergo elongation, a critical step of gastrulation. We recreated the system in HCT-116 cells, and we show that the loss of Sin3A was marked by a loss of proliferative potential.