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

描述（由申请人提供）：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，但越来越明显的是，表观遗传变化在调节癌症发展中起着关键作用。多梳抑制复合物（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EZH 2是致癌PRC 2成员，通过在赖氨酸27处使组蛋白H3三甲基化来启动转录抑制。我们已经证明EZH 2在侵袭性前列腺癌和乳腺癌中过表达，预测疾病结果，并且是癌细胞存活所必需的。此外，我们的研究表明，EZH 2下调多种肿瘤抑制因子，microRNA-101的基因组缺失导致侵袭性肿瘤中EZH 2的表达不受调节。虽然EZH 2在调节蛋白质编码基因中的作用是已知的，但其在调节microRNA（miR）表达中的作用尚未研究。miR是细胞功能的关键调节因子，并且通常在癌症中改变，包括下调几种肿瘤抑制miR。因此，我们假设转录抑制因子EZH 2通过表观遗传沉默在调节microRNA表达中起关键作用，并且EZH 2调节的miR在PCa进展中起关键作用。 我们的初步数据表明，EZH 2下调多种miR，包括miR-203和miR-200 a，miR-200 bc家族。miR-203和mir-200 a，bc依次调节PRC 1成员、BMI 1和RING 2。因此，本提案的目的是扩展这些发现，并进一步了解EZH 2对miR的调节以及这些miR在PCa发展中的作用。为了实现这些目标，在具体目标1中，我们将研究EZH 2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前列腺细胞系的RNA进行miR分析，其中EZH 2表达被调节。我们将验证EZH 2调控的miR在前列腺肿瘤组织中的表达，并将其与EZH 2表达相关联。在具体目标2中，我们将研究EZH 2调节的miR在PCa中的作用及其在靶向PRC 1成员中的作用。在具体目标3中，我们将研究EZH 2调节的miR在前列腺肿瘤发生中的作用。我们将使用选择的EZH 2调节的miR，其直接被EZH 2抑制，并使用PCa的细胞系和体内模型来表征其调节的后果。与公共卫生的相关性：拟议工作的成功完成将为PCa发展中的miR和表观遗传调节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，这反过来可能通过更好的诊断和疾病监测来改善PCa治疗。我们的研究结果可能最终为在癌症中治疗性地“重新引入”EZH 2抑制的肿瘤抑制miR提供证据。

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.