Epigenetic profiling of EMT in pancreatic cancer cells (ChIP-seq)

Epithelial plasticity – reversible modulation of a cell’s epithelial and mesenchymal features – is associated with cancer metastasis and chemoresistance. This process is classically driven by a small cohort of transcription factors, several of which have been reported to functionally interact with epigenetic modifiers at the promoters of key epithelial and mesenchymal genes. However, the extent to which epigenetic changes underlie epithelial plasticity at the genomic level remains largely unknown. Here we show that genome-wide regulation of a single histone mark, H3K36me2, is critical for establishing stable epithelial-mesenchymal states in various genetic, chemical, and cellular contexts. Through targeted CRISPR-Cas9 screening, we discovered two histone-modifying enzymes involved in the writing and erasing of H3K36me2 that act reciprocally to regulate epithelial-mesenchymal identity, tumor differentiation, and metastasis. Using genetic approaches to inhibit H3K36me2, we found that modulation of the mark itself is a conserved mechanism underlying the mesenchymal state. Although H3K36me2 is distributed broadly throughout the genome, it is associated with altered enhancer activity affecting a relatively small number of genes, including those associated with master epithelial-mesenchymal regulatory factors. Our results thus outline an epigenome-scale mechanism by which a specific histone modification regulates cellular plasticity in cancer. 18 samples were sequenced for 4 histone marks (H3K36me2, H3K27ac, H3K4me1, H3K27me3) in duplicate and inputs. Comparison groups include WT and sgNsd2 mouse pancreatic cancer cells from the Panc02 cell line. Sonicated Drosophila melanogaster DNA was spiked into each sample at 1-2.5% of mouse chromatin.

上皮可塑性（epithelial plasticity）——即细胞上皮与间质表型的可逆调控——与癌症转移及化疗耐药密切相关。该过程经典上由一小簇转录因子驱动，已有研究报道其中多个因子可在关键上皮及间质基因的启动子区域，与表观遗传修饰因子（epigenetic modifiers）发生功能性互作。然而，在基因组层面上，表观遗传变化在上皮可塑性形成过程中所扮演的具体作用程度，目前仍知之甚少。本研究证实，单一组蛋白修饰标记（histone mark）H3K36me2的全基因组调控，在多种遗传、化学及细胞环境中，对于建立稳定的上皮-间质细胞状态至关重要。通过靶向CRISPR-Cas9筛选实验，我们发现两种分别负责H3K36me2的写入与移除的组蛋白修饰酶，二者以反向调控的方式共同影响上皮-间质细胞身份、肿瘤分化及转移过程。通过遗传手段抑制H3K36me2的水平后，我们发现该修饰标记本身的调控是维持间质细胞状态的保守机制。尽管H3K36me2广泛分布于全基因组，但它与少数基因的增强子活性改变密切相关，这些基因涵盖核心上皮-间质调控因子相关基因。综上，本研究揭示了一种表观基因组层面的调控机制，即特定组蛋白修饰可调控癌症中的细胞可塑性。本数据集共包含18份样本，针对4种组蛋白修饰标记（H3K36me2、H3K27ac、H3K4me1、H3K27me3）设置了重复样本及输入对照样本并完成测序。比较实验组包括源自Panc02细胞系的野生型（WT）小鼠胰腺癌细胞与sgNsd2基因敲除小鼠胰腺癌细胞。每份样本中均掺入了占小鼠染色质总量1%-2.5%的超声处理黑腹果蝇（Drosophila melanogaster）DNA作为内参。