In situ Hi-C in Smchd1 wild-type and Smchd1 deleted neural stem cells.. In situ Hi-C in Smchd1 wild-type and Smchd1 deleted neural stem cells.

We sought to examine whether the non-canonical SMC protein Smchd1 plays a role in chromosome conformation. We used in situ Hi-C to analyse chromosome conformation changes upon deletion of the epigenetic regulator Smchd1 in female neural stem cells. In parallel, we analysed nucleosome accessibility using ATAC-seq, gene expression using RNA-seq, chromatin marks H3K27me3 and H3K27ac and Ctcf binding using ChIP-seq. We additionally analysed Smchd1 binding genome-wide using ChIP-seq. Together, we find that deletion of Smchd1 alters chromosome conformation at Smchd1 target genes including the inactive X chromosome, Hox genes and imprinted loc. Smchd1 deletion results in gain in Ctcf binding and activation of enhancers. We propose Smchd1 functions by limiting Ctcf-mediated chromosome looping. Overall design: n=3 Smchd1del/del vs n=3 Smchd1fl/fl female neural stem cell lines analysed by in situ Hi-C

本研究旨在探究非典型SMC蛋白Smchd1是否在染色体构象调控中发挥作用。我们采用原位Hi-C（in situ Hi-C）技术，分析雌性神经干细胞中表观遗传调控因子Smchd1敲除后染色体构象的变化。同时，我们利用ATAC-seq检测核小体可及性、RNA-seq分析基因表达水平，通过ChIP-seq（染色质免疫共沉淀测序）检测染色质修饰标记H3K27me3、H3K27ac以及CTCF的结合情况。此外，我们还通过ChIP-seq对Smchd1在全基因组范围内的结合特征进行了分析。综合分析结果显示，Smchd1敲除会改变其靶基因位点的染色体构象，涉及失活X染色体、Hox基因及印记基因座。Smchd1敲除可导致CTCF结合增强并激活增强子。我们推测Smchd1通过抑制CTCF介导的染色体环化发挥调控功能。实验设计：以3株Smchd1del/del雌性神经干细胞系为实验组、3株Smchd1fl/fl雌性神经干细胞系为对照组，采用原位Hi-C技术进行染色体构象分析。