Molecular genetics of Pelizaeus-Merzbacher disease

项目摘要 Pelizaeus-Merzbacher病（PMD）是一种X连锁脑白质营养不良，由 蛋白脂质蛋白1基因（PLP 1），编码主要的中枢神经系统髓鞘蛋白。重复 含有PLP 1的基因区域是PMD的最常见原因，占病例的60%以上。 尽管疾病的严重程度可能存在变异性，即使在一个家庭中，大多数患者 PLP 1重复具有相似的表型，在出生后的最初几个月内出现眼球震颤和张力减退， 以及在头十年期间的肢体和步态共济失调、痉挛性四肢轻瘫以及认知和视觉障碍。 大多数患者的运动里程碑和语言也会延迟。基因的分子机制 PMD临床表型的分子基础也不清楚。在我们以前 对13例PMD患者的重组连接DNA序列进行了研究，我们发现数据 这与导致基因突变的同源、非同源重组机制相一致， 复制品然而，Lupski和他的同事最近通过对重组连接的分析提出， 在2例具有复杂重排的患者中，基于复制的机制也可能参与其中。 过程在本研究中，我们将分析重组断点的结构和序列 在一个更大的基因重复患者队列中，使用了包括高密度 寡核苷酸阵列。此外，为了进一步了解这些引起临床表型的基础， 我们将构建一个PMD小鼠模型，该模型含有一个大的，复杂的重复基因， X染色体的PLP 1区域。已经构建了含有这种复制的ES细胞， 包含X染色体重排部分的基因正在被制造。然后分析动物的 他们的临床表型，表达的基因编码的重复，包括PLP 1，以及 髓鞘形成程序中其他基因的表达。分离的重排部分的X- 还将在女性携带者中检查其对X-失活的影响。综上所述各项 这些研究将为PMD中基因复制的机制提供重要的新信息， 这种基因复制对基因表达和神经功能的影响。我们组特别好 准备进行这些研究，因为我们已经开发了独特的患者资源和专业知识， 在过去的十年里，我们从PMD诊断和研究的承诺中获益匪浅。具体 目的是：（1）验证PMD患者基因组重排频繁发生的假设， 重排，包括第二个重复区域和缺失、三重或倒置区域，一致 具有偶联的同源、非同源重组机制。(2)为了验证这个假设， 在Plp 1基因座存在基因重复改变Plp 1基因表达，导致髓鞘破坏 在男性和女性的X染色体失活模式的补偿性偏斜的程序。

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.