Ademar Jesus Silva

Endogenous and exogenous immunoprecipitations Harvested cells were washed twice with cold PBS and washed pellets resuspended with Buffer A (25mM HEPES pH 7.6, 5mM MgCl2, 25mM KCl, 0.05mM EDTA, 10% (v/v) glycerol, 0.1% (v/v) NP40, 1mM DTT and 1mM PMSF) and kept on ice for 10 minutes. Samples were centrifuged at 1,500rpm for 10 minutes at 4°C and the supernatant was discarded.

Abstract Diffuse midline glioma (DMG) is a fatal childhood brain tumour characterised primarily by mutant histone H3 (H3K27M). H3K27M causes a global reduction in Polycomb Repressive Complex 2 (PRC2)-mediated H3K27me3 by inhibiting PRC2 enzymatic activity. Paradoxically, PRC2 is essential in DMG tumour cells where residual complex activity is required for oncogenic gene repression, although the molecular mechanisms acting downstream of PRC2 in this context are poorly understood.