DE PEDIATRIC COBRE: MOLECULAR MECHANISMS IN PELIZAEUS MERZBACHER DISEASE

DE PEDIATRIC COBRE：Pelizaeus Merzbacher 病的分子机制

• 批准号: 7720951
• 负责人: Grace M. Hobson
• 金额: $ 14.35万
• 依托单位: ALFRED I. DU PONT HOSP FOR CHILDREN
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-01 至 2009-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7720951
• 关键词: 5' Splice Site; Accounting; Affect; Alternative Splicing; Biological Assay; Childhood; Computer Retrieval of Information on Scientific Projects Database; Computer Simulation; Coupled; Defect; Diagnostic Procedure; Disease; ES Cell Line; Engineering; Enhancers; Exons; Family; Funding; Gene Expression; Gene Transfer; Genes; Genomics; Grant; Hereditary Disease; Institution; Introns; Link; Location; Molecular; Molecular Cytogenetics; Mus; Mutation; Myelin Proteins; Oligodendroglia; Pathogenesis; Pelizaeus-Merzbacher Disease; Play; Point Mutation; Protein Binding; Proteins; Proteolipids; RNA Splicing; Relative (related person); Research; Research Personnel; Resources; Role; Source; Structure; Testing; Trans-Activators; Transfer RNA; United States National Institutes of Health; cis acting element; disease phenotype; dosage; improved; leukodystrophy; novel; size

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Pelizaeus-Merzbacher disease (PMD), an X-linked leukodystrophy, is caused by defects of the proteolipid protein 1 gene (PLP1) that encodes the major CNS myelin protein. Approximately 60% of families have a genomic duplication that includes the PLP1 gene, and 15-20% of families have PLP1 point mutations. Studies in mice have shown that an increased dosage of PLP1 can account for disease pathogenesis. Further, studies have shown that mutations in noncoding regions can alter PLP1 expression levels or the ratio of the alternatively spliced forms PLP1 and DM20. Our long-term objective is to understand the molecular mechanisms involved in generating the PMD phenotype so rational treatments and improved diagnostic techniques can be developed. In the first aim, results on location, size, structure and sequence at the junctions of PMD duplications support the hypothesis that most are caused by a novel coupled homologous and nonhomologous mechanism. We will continue a combined cytogenetic, molecular, and in silico approach to refine our understanding of the mechanism. In the second aim, the hypothesis that the duplication structure affects expression of PLP1 is being tested by engineering different duplications into ES cell lines and analyzing the structural effects on gene expression before and after differentiation into oligodendrocytes. In the third aim, the hypothesis that some PMD mutations dysregulate splicing of PLP1 is being tested by using gene transfer and RNA-protein binding assays to investigate cis-acting elements and trans-acting factors involved in PLP1/DM20 alternative splicing. We have determined that exon and intron splicing enhancers and the relative strength of the PLP1 and DM20 donor splice sites play an important role in PLP1 alternative splicing. The results of these studies have greatest impact potential through their generalizability to other genomic and splicing diseases, which have only recently been recognized as important types of genetic disease.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 Pelizaeus-Merzbacher 病 (PMD) 是一种 X 连锁脑白质营养不良，是由编码主要 CNS 髓磷脂蛋白的蛋白脂质蛋白 1 基因 (PLP1) 缺陷引起的。大约 60% 的家族存在包含 PLP1 基因的基因组重复，15-20% 的家族存在 PLP1 点突变。对小鼠的研究表明，增加 PLP1 剂量可以解释疾病的发病机制。此外，研究表明非编码区的突变可以改变 PLP1 表达水平或选择性剪接形式 PLP1 和 DM20 的比例。我们的长期目标是了解产生 PMD 表型的分子机制，以便开发合理的治疗方法和改进的诊断技术。第一个目标是，关于 PMD 重复连接处的位置、大小、结构和序列的结果支持这样的假设：大多数重复是由一种新颖的耦合同源和非同源机制引起的。我们将继续结合细胞遗传学、分子学和计算机方法来完善我们对该机制的理解。在第二个目标中，通过将不同的重复工程设计到 ES 细胞系中并分析分化为少突胶质细胞之前和之后对基因表达的结构影响，正在测试重复结构影响 PLP1 表达的假设。第三个目标是，通过使用基因转移和 RNA 蛋白结合测定来研究 PLP1/DM20 选择性剪接中涉及的顺式作用元件和反式作用因子，正在测试一些 PMD 突变使 PLP1 剪接失调的假设。我们已经确定外显子和内含子剪接增强子以及 PLP1 和 DM20 供体剪接位点的相对强度在 PLP1 选择性剪接中发挥重要作用。这些研究的结果通过其对其他基因组和剪接疾病的普遍适用性而具有最大的影响潜力，这些疾病最近才被认为是重要的遗传病类型。