Higher-order chromatin organization defines PR and PAX2 binding to regulate endometrial cancer cell gene expression (ATACseq)

Estrogen (E2) and Progesterone (Pg) via their specific receptors, ER and PR respectively, are major determinants in the development and progression of endometrial malignancies. We have studied how E2 and the synthetic progestin R5020 affect genomic function in Ishikawa endometrial cancer cells. Using ChIPseq in cells exposed to the corresponding hormones, we identified cell specific binding sites for ER (ERbs) and PR (PRbs), mostly binding to independent sites and both adjacent to PAXbs. Long-range interactions (HiC) showed enrichment of PRbs and PAXbs, which we call progestin control regions (PgCRs) inside TADs with differentially progestin-regulated genes. Effects of hormone treatments on gene expression were detected by RNAseq. PgCRs correlate with open chromatin independently of hormonal stimuli. In summary, endometrial response to progestins in differentiated endometrial tumor cells results in part from binding of PR to compartmentalized PgCRs in hormone-independent open chromatin, which include binding of partner transcription factors, in particular PAX2. Endometrial adenocarcinoma Ishikawa cells were treated with R5020 10nM or E2 10nM for different times and submitted to ATACseq.

雌二醇（Estrogen, E2）与孕酮（Progesterone, Pg）可分别通过其特异性受体雌激素受体（ER）与孕激素受体（PR），成为子宫内膜恶性肿瘤发生与进展的关键调控因子。本研究以石川（Ishikawa）子宫内膜癌细胞系为模型，探究了E2与合成孕激素R5020对其基因组功能的影响。研究人员利用染色质免疫共沉淀测序（ChIP-seq），在经对应激素处理的细胞中鉴定出雌激素受体与孕激素受体的细胞特异性结合位点（分别记为ERbs与PRbs），二者多数独立结合，且均紧邻PAX结合位点（PAXbs）。染色质长距离相互作用分析（Hi-C）结果显示，在携带孕激素差异调控基因的拓扑关联结构域（TADs）内，PRbs与PAXbs存在富集现象，我们将此类区域命名为孕激素调控区域（PgCRs）。通过RNA测序（RNA-seq）检测到激素处理对基因表达的调控效应。PgCRs与不依赖激素刺激的开放染色质区域存在显著关联。综上，分化型子宫内膜肿瘤细胞对孕激素的应答，部分源于PR结合至激素非依赖型开放染色质中的区域化PgCRs，此类区域包含伴侣转录因子（尤其是PAX2）的结合位点。本研究中，子宫内膜腺癌细胞系Ishikawa经10nM R5020或10nM E2处理不同时长后，被用于开展转座酶可及性染色质测序（ATAC-seq）。