Regulation of Polycomb Repressive Complex 1 by EZH2 Regulated microRNAs in Cancer

描述（申请人提供）：前列腺癌（PCa）是美国男性癌症相关死亡的第二大常见原因。虽然多种分子事件有助于前列腺癌的进展，但表观遗传变化在调节癌症发展中起着关键作用已经越来越明显。多梳抑制复合体（PRC）成员通过表观遗传修饰组蛋白来维持细胞的基因表达状态。组蛋白甲基转移酶EZH2，一个致癌的PRC2成员，通过在赖氨酸27处三甲基化组蛋白H3来启动转录抑制。我们已经证明EZH2在侵袭性前列腺癌和乳腺癌中过度表达，预测疾病结局，并且是癌细胞存活所必需的。此外，我们的研究表明EZH2下调多种肿瘤抑制因子，microRNA-101的基因组缺失导致EZH2在侵袭性肿瘤中的不调节表达。虽然已知EZH2在调节蛋白编码基因中的作用，但其在调节microRNA （miR）表达中的作用尚未被研究。MiRs是细胞功能的关键调节因子，在癌症中通常发生改变，包括下调几种肿瘤抑制MiRs。因此，我们假设转录抑制因子EZH2通过表观遗传沉默在调节microRNA表达中起关键作用，EZH2调节的miRs在PCa的进展中起关键作用。我们的初步数据表明EZH2下调了多个mir，包括miR-203和miR-200a、miR-200bc家族。MiR-203和mir-200a， bc反过来调控PRC1成员BMI1和RING2。因此，本提案的目的是扩展这些发现，并进一步了解EZH2对miRs的调节以及这些miRs在PCa发展中的作用。为了实现这些目标，在Specific Aim 1中，我们将研究EZH2在多种细胞类型中调节miR表达的作用。我们将首先使用来自EZH2表达被调节的前列腺细胞系的RNA进行miR分析。我们将验证EZH2在前列腺肿瘤组织中调节miR表达，并将其与EZH2表达联系起来。在Specific Aim 2中，我们将研究ezh2调控的miRs在PCa中的作用以及它们在靶向PRC1成员中的作用。在Specific Aim 3中，我们将研究ezh2调控的miRs在前列腺肿瘤发生中的作用。我们将使用EZH2直接抑制的EZH2调节的miRs，并使用细胞系和体内PCa模型表征其调节的后果。与公共卫生相关：成功完成拟议的工作将为前列腺癌发展中miR和表观遗传调控因子的复杂网络提供证据，并确定潜在的诊断和预后标志物，从而通过更好的诊断和疾病监测改善前列腺癌治疗。我们的结果可能最终为治疗“重新引入”ezh2抑制的肿瘤抑制miRs在癌症中提供依据。

DESCRIPTION (provided by applicant): Prostate cancer (PCa) is the second most common cause of cancer-related death in men in the United States. While multiple molecular events contribute to prostate cancer progression, it has become increasingly evident that epigenetic changes play pivotal roles in regulating cancer development. Polycomb repressive complex (PRC) members maintain the gene expression status of a cell by epigenetically modifying histone proteins. Histone methyltransferase EZH2, an oncogenic PRC2 member, initiates the transcriptional repression by trimethylating histone H3 at lysine 27. We have shown that EZH2 is overexpressed in aggressive prostate and breast cancers, predicts disease outcome, and is required for cancer cell survival. In addition, our studies have shown that EZH2 down-regulates multiple tumor suppressors and a genomic loss of microRNA-101 results in unregulated expression of EZH2 in aggressive tumors. While the role of EZH2 in regulating protein-coding genes is known, its role in regulating microRNA (miR) expression has not been studied. MiRs are critical regulators of cellular functions and are commonly altered in cancers including down regulation of several tumor suppressor miRs. Thus, we hypothesize that transcriptional repressor EZH2 plays a key role in regulating microRNA expression by epigenetic silencing and EZH2-regulated miRs play critical roles in PCa progression. Our preliminary data suggest that EZH2 down regulates multiple miRs, including miR-203 and the miR-200a, miR-200bc family. MiR-203 and mir-200a, bc in turn regulate PRC1 members, BMI1 and RING2. Thus the aims of this proposal are to extend these findings and gain further insights into the regulation of miRs by EZH2 and the role of these miRs in PCa development. In order to accomplish these goals, in Specific Aim 1, we will investigate the role of EZH2 in regulating miR expression in multiple cell types. We will first perform miR profiling using RNA from prostate cell lines in which EZH2 expression is modulated. We will validate the EZH2 regulated miR expression in prostate tumor tissues and correlate them with EZH2 expression. In Specific Aim 2, we will investigate the role of EZH2-regulated miRs in PCa and their role in targeting the PRC1 members. In Specific Aim 3, we will investigate the role of EZH2-regulated miRs in prostate tumorigenesis. We will use select EZH2-regulated miRs that are directly repressed by EZH2 and characterize the consequences of their modulation using both cell line and in vivo models of PCa. Relevance to Public Health: Successful completion of the proposed work will provide evidence for an intricate network of miR and epigenetic regulators in PCa development and identify potential diagnostic and prognostic markers, which in turn may improve PCa therapy through better diagnosis and disease monitoring. Our results may ultimately provide credence for therapeutic "re-introduction" of EZH2-repressed tumor suppressor miRs in cancer.