3D genome organization in the epithelial-mesenchymal transition spectrum [scHi-C_PEO1HEYA8]

Background The plasticity along the epithelial-mesenchymal transition (EMT) spectrum has been shown to be regulated by various epigenetic repertoires. Emerging evidence of local chromatin conformation changes suggests that regulation of EMT may occur at a higher order of three-dimensional genome level. Results We perform Hi-C analysis and combine ChIP-seq data across cancer cell lines representing different EMT states. We demonstrate that the epithelial and mesenchymal genes are regulated distinctively. We find that EMT genes are regulated within their topologically associated domains (TADs), with only a subset of mesenchymal genes being influenced by A/B compartment switches, indicating topological remodelling is required in the transcriptional regulation of these genes. At the TAD level, epithelial and mesenchymal genes are associated with different regulatory trajectories. The epithelial gene-residing TADs are enriched with H3K27me3 marks in the mesenchymal-like states. The mesenchymal gene-residing TADs , which do not show enrichment of H3K27me3 in epithelial-like states, exhibit increased interaction frequencies with regulatory elements in the mesenchymal-like states. Conclusions We propose a novel workflow coupling immunofluorescence and dielectrophoresis to unravel EMT heterogeneity at single-cell resolution. The predicted three-dimensional structures of chromosome 10, harboring Vimentin, identify cell clusters of different states. Our results pioneer a novel avenue to decipher the complexities underlying the regulation of EMT and may infer the barriers of plasticity in the 3D genome context. single-cell Hi-C seq profiling of ovarian cancer cell line PEO1 and HEYA8.

**研究背景**
上皮间质转化（Epithelial-Mesenchymal Transition, EMT）谱式中的细胞可塑性已被证实受多种表观遗传调控机制的调控。近期关于局部染色质构象变化的研究证据表明，EMT的调控可能发生在更高维度的三维基因组层面。

**研究结果**
本研究针对呈现不同EMT状态的癌细胞系开展Hi-C测序分析，并整合染色质免疫共沉淀测序（Chromatin Immunoprecipitation sequencing, ChIP-seq）数据。研究证实，上皮细胞与间质细胞的基因调控模式存在显著差异：EMT相关基因的调控发生在其自身的拓扑关联结构域（Topologically Associated Domains, TADs）内，仅部分间质基因受A/B区室切换的影响，这提示拓扑结构重塑是此类基因转录调控的必要条件。在TAD层面，上皮与间质相关基因呈现出不同的调控轨迹：在上皮样状态下，携带上皮基因的TAD富集H3K27me3修饰标记；而携带间质基因的TAD在上皮样状态下未出现H3K27me3富集，但在间质样状态下与调控元件的相互作用频率显著升高。

**研究结论**
本研究提出了一种结合免疫荧光与介电电泳的全新实验流程，可实现单细胞分辨率下EMT异质性的解析。针对携带波形蛋白（Vimentin）的10号染色体构建的预测三维结构，可区分不同状态的细胞簇。本研究为解析EMT调控的复杂机制开辟了新途径，或可帮助阐释三维基因组语境下细胞可塑性的调控障碍。本数据集包含卵巢癌细胞系PEO1与HEYA8的单细胞Hi-C测序分析数据。