Examining the role of ATRX-RNA interactions in ATRX and PRC2 localization and gene expression.. Examining the role of ATRX-RNA interactions in ATRX and PRC2 localization and gene expression.

We disrupted the RNA binding region in endogenous ATRX and tested localization of ATRX, EZH2 and H3K27me3 by CUT&RUN. We show that ATRX-RNA binding is important to regulate ATRX and PRC2 localization to specific genes. Overall design: We performed CUT&RUN for ATRX, EZH2 and H3K27me3 in WT and ATRX△RBR (ATRX with RNA binding region deleted) using the protocol described in Skene et al, Nature protocols, 2018). RNA Seq was performed in WT and ATRX△RBR.

我们对内源ATRX的RNA结合区域实施了敲除，并通过CUT&RUN（靶位点切割释放测序技术）检测了ATRX、EZH2及H3K27me3的定位情况。本研究证实，ATRX与RNA的结合对于调控ATRX以及多梳抑制复合体2（PRC2, Polycomb Repressive Complex 2）在特定基因上的定位具有关键作用。
整体实验设计：我们参照Skene等人2018年发表于《Nature Protocols》的实验方案，分别在野生型（WT, Wild Type）和ATRXΔRBR（RNA结合区域缺失的ATRX）细胞中，针对ATRX、EZH2及H3K27me3开展了CUT&RUN实验；同时在上述两种细胞系中完成了转录组测序（RNA Seq, RNA Sequencing）。