Molecular genetics of Pelizaeus-Merzbacher disease

项目总结 Pelizaeus-Merzbacher病(PMD)是一种X连锁脑白质营养不良症，由脑白质细胞的遗传缺陷引起。 蛋白脂蛋白1基因(PLP1)，编码主要的中枢神经系统髓鞘蛋白。复制 含有PLP1的基因区域是PMD最常见的原因，占PMD的60%以上。 尽管疾病的严重程度可能会有所不同，但即使在一个家庭内，大多数患有 PLP1基因重复与出生后最初几个月出现眼球震颤和低眼压的表型相似， 以及肢体和步态共济失调、痉挛四肢瘫痪，以及第一个十年的认知和视觉障碍。 大多数患者的运动里程碑和语音也会延迟。既不是基因的分子机制 PMD的临床表型的分子基础也不是很清楚。在我们之前的 对13例PMD患者的重组连接DNA序列的研究，我们发现了数据 与导致该基因的耦合的同源和非同源重组机制一致 复制品。然而，卢普斯基和他的同事最近从对重组连接的分析中提出 在2名有复杂重排的患者中，基于复制的机制可能也参与了这一过程 进程。在拟议的研究中，我们将分析重组断点的结构和序列 在使用多种策略组合的基因复制患者中，包括高密度 寡核苷酸阵列。此外，为了进一步了解这些疾病引起的临床表型的基础 复杂的基因复制，我们将构建一种PMD的小鼠模型，该模型包含一个大型的、复杂的复制的 X染色体的PLP1区域。含有这种复制的ES细胞已经被构建，小鼠 包含X染色体重排部分的基因正在制造中。然后将对动物进行分析 他们的临床表型，复制编码的基因的表达，包括PLP1，以及 其他基因在髓鞘形成过程中的表达。X-的重排部分的分离 还将对女性携带者的染色体进行检查，以确定其对X-染色体失活的影响。这些加在一起， 研究将提供有关PMD基因复制机制的重要新信息，以及 该基因复制对基因表达和神经功能的影响。我们这群人特别好 准备进行这些研究，因为我们开发了独特的患者资源和专业知识 在过去十年中，我们从对PMD诊断和研究的承诺中获益。具体的 目的是：(1)检验PMD患者基因组重排经常以复杂形式发生的假设 重排，包括第二个重复区域和删除、三重或倒置区域，一致 具有耦合的同源和非同源重组机制。(2)检验假设 Plp1基因重复改变Plp1基因表达导致髓鞘破坏 男性的程序和女性的X染色体失活模式的补偿性倾斜。

PROJECT SUMMARY Pelizaeus-Merzbacher disease (PMD) is an X-linked leukodystrophy caused by genetic defects of the proteolipid protein 1 gene (PLP1) that encodes the major central nervous system myelin protein. Duplication of a gene region containing PLP1 is the most common cause of PMD, accounting for over 60% of cases. Although there can be variability in the severity of the disease, even within a single family, most patients with PLP1 duplications have a similar phenotype with onset of nystagmus and hypotonia in the first months of life, and limb and gait ataxia, spastic quadriparesis, and cognitive and visual impairment during the first decade. Motor milestones and speech are also delayed in most patients. Neither the molecular mechanisms of gene duplication nor the molecular basis for the clinical phenotype in PMD is well understood. In our previous studies of the DNA sequences of recombinant junctions in 13 patients with PMD, we have found data consistent with a coupled homologous, nonhomologous recombination mechanism causing the gene duplications. Lupski and co-workers, however, recently suggested from the analysis of recombinant junctions in 2 patients with complex rearrangements that a replication-based mechanism may also be involved in this process. In the proposed studies, we will analyze the structure and sequence of the recombination breakpoints in a larger cohort of patients with gene duplications using a combination of strategies, including high-density oligonucleotide arrays. In addition, to further understand the basis of the clinical phenotype caused by these complex gene duplications, we will construct a mouse model of PMD containing a large, complex duplication of the PLP1 region of the X-chromosome. ES cells containing this duplication have been constructed, and mice containing the rearranged portion of the X-chromosome are being made. Animals will then be analyzed for their clinical phenotype, the expression of genes encoded by the duplication, including PLP1, as well as expression of other genes in the program of myelination. Segregation of the rearranged portion of the X- chromosome will also be examined in female carriers for its effects on X-inactivation. Taken together, these studies will provide important new information on the mechanism of gene duplication in PMD, as well as the effect of this gene duplication on gene expression and neurological function. Our group is particularly well poised to perform these studies because of the unique patient resource we have developed and the expertise we have gained from a commitment to PMD diagnostics and research during the past decade. The specific aims are: (1) To test the hypothesis that genomic rearrangements in PMD patients frequently occur as complex rearrangements, including a second duplicated region and deleted, triplicated or inverted regions, consistent with a coupled homologous, nonhomologous recombination mechanism. (2) To test the hypothesis that the presence of a gene duplication at the Plp1 locus alters Plp1 gene expression leading to disruption of the myelin program in males and compensatory skewing of the X-chromosome inactivation pattern in females.