DNA methylation in normal versus malignant melanocytes

正常黑素细胞与恶性黑素细胞中的 DNA 甲基化

基本信息

批准号：
6952686
负责人：
SHERMAN Morton WEISSMAN
金额：
$ 8.18万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-09-25 至 2007-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6952686
关键词：
CpG islands DNA methylation chromatin immunoprecipitation functional /structural genomics gene expression profiling genetic markers high throughput technology melanocyte microarray technology neoplasm /cancer genetics neoplastic cell neoplastic growth nucleic acid probes

项目摘要

DESCRIPTION (provided by applicant): Aberrant DNA methylation at CpG dinucleotide islands, the major epigenetic modification of mammalian genomes, is frequent during tumor progression. For example, inactivation by DNA methylation is the cause for down-regulation of tumor suppressors, apoptotic factors, DNA repair enzymes, and adhesion molecules involved in malignant progression to melanoma and other cancers. In addition, a panel of melanoma antigens, known as cancer/testis-antigens is aberrantly re-expressed due to DNA demethylation of CpG promoter islands. In this project normal human melanocytes and primary melanoma cells will be employed to assess changes in CpG methylation patterns and the genes they affect in a high throughput manner. This will be accomplished by probing promoter chip arrays and genomic tiling arrays employing modified DNA and ChIP on chip procedures. The information will be used to assess CpG island methylation as a marker for tumor progression of early melanocytic lesion. The specific aims are: Aim 1: To establish a procedure to probe promoter chip and chromosome tiling arrays for analysis of the methylation status of CpG rich islands on a genome-wide basis, determine the methylation status of CpG island regions in normal melanocytes versus primary melanoma cells. Aim 2: To employ chromatin immunoprecipitation procedure to identify binding regions of methyl-CpG-binding domain proteins (MBP) in the promoter region of these genes (Chip/chip). Aim 3: To validate the results of these two procedures by the bisulfite DNA modification method combined with sequencing of specific DNA regions and gene expression analysis. The information derived from these experiments will reveal the identity of a large fraction of genes whose expression is modulated by CpG island methylation/demethylation events and the extent of changes in CpG island methylation in the early progression of melanocytes to melanomas. The data will then be used to design a simpler and cheaper CpG island microarray chip that can be sed to probe the status of DNA methylation in melanocytic lesions as markers for malignancy.

描述（由申请人提供）：CpG 二核苷酸岛的异常 DNA 甲基化是哺乳动物基因组的主要表观遗传修饰，在肿瘤进展过程中很常见。例如，DNA甲基化导致的失活是肿瘤抑制因子、凋亡因子、DNA修复酶和粘附分子下调的原因，这些分子参与黑色素瘤和其他癌症的恶性进展。此外，由于 CpG 启动子岛的 DNA 去甲基化，一组黑色素瘤抗原（称为癌症/睾丸抗原）会异常重新表达。在这个项目中，正常人类黑色素细胞和原代黑色素瘤细胞将用于以高通量方式评估 CpG 甲基化模式及其影响的基因的变化。这将通过使用改良的 DNA 和芯片上 ChIP 程序探测启动子芯片阵列和基因组平铺阵列来完成。该信息将用于评估 CpG 岛甲基化作为早期黑素细胞病变肿瘤进展的标志物。具体目标是：目标 1：建立探测启动子芯片和染色体平铺阵列的程序，用于在全基因组基础上分析富含 CpG 岛的甲基化状态，确定正常黑色素细胞与原发性黑色素瘤细胞中 CpG 岛区域的甲基化状态。目标 2：采用染色质免疫沉淀程序来鉴定这些基因启动子区域中甲基 CpG 结合域蛋白 (MBP) 的结合区域（芯片/芯片）。目标 3：通过亚硫酸氢盐 DNA 修饰方法结合特定 DNA 区域测序和基因表达分析来验证这两个程序的结果。从这些实验中获得的信息将揭示其表达受 CpG 岛甲基化/去甲基化事件调节的大部分基因的身份，以及黑色素细胞向黑色素瘤早期进展中 CpG 岛甲基化的变化程度。然后，这些数据将用于设计一种更简单、更便宜的 CpG 岛微阵列芯片，可用于探测黑素细胞病变中 DNA 甲基化的状态，作为恶性肿瘤的标记。