Project description:Total RNA was extracted using TRIzol from PEO4 and OVCAR-3 cells treated for 72 hours with 10uM GSK6853 or DMSO as a control. In case of BRPF1 silencing experiment, RNA extraction was performed from cells transfected with BRPF-targeting siRNA or nontarget (scramble) siRNA 96 hours post-transfection using TRIzol and Zymo RNA Clean & Concentrator-5 columns. Indexed libraries were prepared starting from 1000 ng of total RNA according to Illumina Stranded Total RNA prep Ligation with Ribo-Zero Plus kit. Final libraries were sequenced at a concentration of 0,6 pM/lane on the Illumina Novaseq 6000 using S4 flowcell and v1.5 reagents. The current study was focused on the investigation of molecular mechanisms underlying the antiproliferative effect of BRPF1 inhibition or silencing in ovarian cancer. To this end, transcriptome changes induced by GSK6853 treatment in chemotherapy-resistant PEO4 and OVCAR-3 cells and BRPF1 silencing in OVCAR-3 cells were analysed.
Project description:To determine if an endogenous 22G siRNA sensor transgene is subject to siRNA amplification, small RNAs were deep sequenced from the sensor and from a control transgene that is identical to the sensor but lacks an siRNA target site.
Project description:Post-translational modifications (PTMs) on histones play essential roles in cell fate decisions during development. However, how these PTMs are recognized and coordinated remains to be fully illuminated. Here, we show that BRPF1, a multi-histone binding module protein, is essential to maintain pluripotency in human embryonic stem cells (ESCs). BRPF1, H3K4me3 and H3K23ac substantially co-occupy the active and stemness genes in hESCs. BRPF1 deletion impairs H3K23ac and leads to pluripotency exit and closed chromatin accessibility on stemness genes. Deletion of the N terminal or PHD-zinc knuckle-PHD (PZP) modules completely impairs BRPF1 to maintain hESC pluripotency while PWWP module deletion only partially impact its functions. Together, we reveal that the multi-histone binding module protein, BRPF1 co-ordinates the crosstalk between different histone modifications to maintain the pluripotency in hESCs.
Project description:Post-translational modifications (PTMs) on histones play essential roles in cell fate decisions during development. However, how these PTMs are recognized and coordinated remains to be fully illuminated. Here, we show that BRPF1, a multi-histone binding module protein, is essential to maintain pluripotency in human embryonic stem cells (ESCs). BRPF1, H3K4me3 and H3K23ac substantially co-occupy the active and stemness genes in hESCs. BRPF1 deletion impairs H3K23ac and leads to pluripotency exit and closed chromatin accessibility on stemness genes. Deletion of the N terminal or PHD-zinc knuckle-PHD (PZP) modules completely impairs BRPF1 to maintain hESC pluripotency while PWWP module deletion only partially impact its functions. Together, we reveal that the multi-histone binding module protein, BRPF1 co-ordinates the crosstalk between different histone modifications to maintain the pluripotency in hESCs.
