Epigenetic regulation of chromosomal instability by EZH2 methyltransferase [RNA-seq]

Chromosomal instability (CIN) and epigenetic alterations, major drivers of tumor evolution, contribute to breast cancer metastasis. Whether CIN is epigenetically regulated in metastatic cells is poorly understood. Here we show that EZH2 expression is highly correlated with genomic copy number alterations across human breast tumors. Consistently, increased expression of EZH2 histone methyl transferase is associated with elevated CIN in triple negative breast cancer (TNBC) metastasis initiating cells. Genetic or pharmacological inhibition of EZH2 suppresses CIN whereas forced overexpression of EZH2 in otherwise non-transformed cells promotes chromosome segregation errors. Integrated global chromatin and transcriptome profiling identified Tankyrase (TNKS), a multifunctional poly (ADP ribose) polymerase (PARP), as a direct target of EZH2 transcriptional suppressive activity. Strikingly, depletion of TNKS was sufficient to abrogate the ability of EZH2 inhibition to suppress CIN. EZH2-mediated suppression of TNKS induced centrosome amplification and multipolar mitosis which often clustered into pseudo-bipolar mitotic spindles exhibiting chromosome segregation defects. Our work establishes a previously unappreciated epigenetic regulatory mechanisms of CIN and suggests that CIN suppression might be an important component of epigenetic modifying therapies such as EZH2 inhibitors which are currently in clinical use. Overall design: RNA sequencing of sorted EO771.ML1 EZH2 high and EZH2 low cells are treated with either DMSO or 5uM EZH2 inhibitor for 7 days.

染色体不稳定性（Chromosomal instability, CIN）与表观遗传改变是驱动肿瘤演进的核心因素，可促进乳腺癌转移。目前学界对转移性肿瘤细胞中CIN是否受表观遗传调控的认知仍十分有限。本研究发现，在人类乳腺肿瘤队列中，EZH2的表达水平与基因组拷贝数变异显著相关。与之相一致的是，三阴性乳腺癌（triple negative breast cancer, TNBC）转移起始细胞中，EZH2组蛋白甲基转移酶的表达上调与CIN水平升高密切相关。通过遗传或药理学手段抑制EZH2可有效抑制CIN，而在非转化细胞中强制过表达EZH2则会加剧染色体分离错误。本研究通过整合全染色质与转录组谱分析，鉴定出多功能多聚ADP核糖聚合酶（poly (ADP ribose) polymerase, PARP）家族成员Tankyrase（TNKS）是EZH2转录抑制活性的直接靶标。值得注意的是，单独敲低TNKS即可完全抵消EZH2抑制对CIN的抑制作用。EZH2介导的TNKS抑制会诱导中心体扩增与多极有丝分裂，这类多极纺锤体常聚类为假双极有丝分裂纺锤体，进而引发染色体分离缺陷。本研究揭示了此前未被阐明的CIN表观遗传调控机制，并提示抑制CIN或许是EZH2抑制剂这类表观遗传修饰疗法的重要作用组分，目前此类抑制剂已进入临床应用。实验整体设计：将分选得到的EO771.ML1细胞分为EZH2高表达亚群与EZH2低表达亚群，分别用二甲基亚砜（DMSO）或5μM的EZH2抑制剂处理7天，随后进行RNA测序。