Analysis of Imprinting Control of H19/Igf2 and Kcnq1

H19/Igf2 和 Kcnq1 的印记控制分析

• 批准号: 7342739
• 负责人: NORA I ENGEL
• 金额: $ 6.66万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7342739
• 关键词: Analysis; Imprinting; Control; H19; Igf2

DESCRIPTION (provided by applicant): The study of epigenetic mechanisms will lead to understanding of how the genome functions as a developmental blueprint and how perturbations of gene expression patterns can lead to cancer. The goal of this proposal is to elucidate epigenetic mechanisms leading to allele-specific gene silencing at two clusters of imprinted genes. Imprinted genes have parental-specific monoallelic expression. Differential DNA methylation at specific sequences is the epigenetic modification most consistently associated with imprinted genes. Many imprinted domains also exhibit expression of non-coding RNAs. Very little is known of the molecular mechanisms involved in allele-specific silencing and the role of the non-coding RNAs in modulating this process. Using the mouse as a model, this proposal will first elucidate the physical interactions between regulatory elements at the H19/lgf2 locus through chromosome conformation capture technology, testing the hypothesis that there are allele-specific interactions. Mice with targeted mutations in the differentially methylated domain (DMD) at H19 will be compared to the wild-type mice. Second, a targeting experiment will test whether transcription of Kcnq1ot, an imprinted antisense non-coding RNA produced from exon 10 of Kcnq1, is required to maintain the imprinting at this locus by inserting a polyadenylation site in the Kcnq1ot1 gene. A third experiment will exploit transgenic RNA interference to target the Kcnq1ot1 transcript and test whether the Kcnq1ot1 RNA itself plays a role in establishing and maintaining allele-specific gene silencing. Genes in both the H19/lgf2 and the Kcnql domains are involved in Beckwith-Wiedemann syndrome, which causes prenatal overgrowth and predisposition to cancer. Loss of imprinting at both domains has also been implicated in several human neoplasias. Thus, these studies will provide a more thorough understanding of the regulatory mechanisms deployed in gene silencing and will give insight into how these mechanisms can go awry in cancerous cells.

描述（由申请人提供）：对表观遗传机制的研究将导致理解基因组作为发育蓝图的功能以及基因表达模式的扰动如何导致癌症。该提案的目的是阐明在两个烙印基因簇下导致等位基因特异性基因沉默的表观遗传机制。印迹基因具有父母特异性的单相表达。在特定序列处的差异DNA甲基化是与印迹基因最一致的表观遗传修饰。许多印迹域也表现出非编码RNA的表达。对于参与等位基因特异性沉默涉及的分子机制以及非编码RNA在调节此过程中的作用，知之甚少。该建议将使用小鼠作为模型，首先将通过染色体构象捕获技术阐明H19/LGF2基因座的调节元素之间的物理相互作用，从而检验了存在等位基因特异性相互作用的假设。将H19的差异甲基化结构域（DMD）中靶向突变的小鼠与野生型小鼠进行比较。其次，一个靶向实验将测试KCNQ1OT的转录是KCNQ1外显子10产生的印记的反义非编码RNA，是否需要通过在KCNQ1OT1基因中插入聚二苯基化位点来维持该基因座的烙印。第三个实验将利用转基因RNA干扰来靶向KCNQ1OT1转录本，并测试KCNQ1OT1 RNA本身是否在建立和维持等位基因特异性基因沉默中起作用。 H19/LGF2和KCNQL结构域中的基因都参与了贝克维斯·韦德曼综合征，这会导致产前过度生长和癌症倾向。在两个领域中失去印迹的丧失也与几种人类肿瘤有关。因此，这些研究将对基因沉默中部署的调节机制有更透彻的了解，并将深入了解这些机制如何在癌细胞中出现问题。