Studies on RNA polymerase III-related leukodystrophy

项目摘要 RNA聚合酶（Pol）III合成大量的非编码RNA，如tRNA、5S rRNA、U6 snRNA 和7SL RNA参与基本细胞过程，包括蛋白质合成，RNA加工和 蛋白质分泌尽管这些RNA在所有真核细胞中都起着重要作用，但Pol III的突变可能会导致 对中枢神经系统（CNS）有显著的选择性作用，如Pol III患者所证明的那样， 相关的脑白质营养不良导致这种常染色体隐性遗传神经退行性疾病的突变已经被发现。 在五种不同的Pol III亚基中发现，疾病发作最常发生在儿童早期。Pol III- 相关的脑白质营养不良是典型的髓鞘生成不足的疾病，尽管患者同时存在一系列的 神经和非神经缺陷。神经缺陷通常包括发育迟缓， 认知和运动功能进行性下降、共济失调和小脑萎缩。非神经功能缺损 可能包括缺牙、近视和内分泌异常。对疾病机制的理解 目前缺乏这些临床特征的基础，治疗仅限于姑息治疗。一 在这两个方面取得进展的重大障碍是缺乏这种疾病的动物模型。 然而，最近的工作已经产生了Pol III相关的脑白质营养不良的小鼠模型， 临床特征的子集。Polr 3a中致病性突变的条件性敲入，其编码最大的 Pol III亚基，特别是在CNS中表达Olig 2的细胞中，导致生长、神经行为和神经行为的减少。 在大脑和脊髓的多个区域中的缺陷和髓鞘形成不足。此外，电子显微镜 沿着对少突胶质细胞群和特异性蛋白标记的分析， 导致疾病发病的不同机制：少突胶质细胞的分化缺陷 前体细胞（OPCs）和成熟少突胶质细胞（OLs）的功能缺陷。然而，分子 发病机制的特征仍然不清楚。为了加深对这方面的了解，长期以来， 这项工作的长期目标是确定Pol III转录组在OPCs和OL中是如何改变的。 突变小鼠，并检查这些变化对蛋白质合成和蛋白质组的影响。 OPCs和OLs以及髓磷脂的脂质体。此外，我们将测试一个模型，用于Pol III的遗传抑制， 相关的脑白质营养不良表型。总之，这些研究将告知如何减少Pol III 转录干扰少突胶质细胞的发生和髓鞘形成，产生重要的机制见解， 可能有益于Pol III相关脑白质营养不良患者的潜在治疗方法。

Project Summary RNA polymerase (Pol) III synthesizes abundant non-coding RNAs such as tRNA, 5S rRNA, U6 snRNA and 7SL RNA that are involved in essential cellular processes including protein synthesis, RNA processing and protein secretion. Despite the essential roles of these RNAs in all eukaryotic cells, mutations in Pol III can have remarkably selective effects on the central nervous system (CNS) as demonstrated by patients with Pol III- related leukodystrophy. Mutations causing this autosomal recessive neurodegenerative disease have been identified in five different Pol III subunits with disease onset occurring most often in early childhood. Pol III- related leukodystrophy is typically a hypomyelinating disorder although patients present with a range of both neurological and non-neurological deficits. Neurological deficits typically include developmental delay with a progressive decline in cognitive and motor function, ataxia and cerebellar atrophy. Non-neurological deficits may include hypodontia, myopia and endocrine abnormalities. An understanding of the disease mechanisms underlying these clinical characteristics is currently lacking and treatments are limited to palliative care. A significant barrier to progress on both these fronts has been the absence of an animal model of the disease. However, recent work has resulted in a mouse model of Pol III-related leukodystrophy that recapitulates a subset of clinical features. Conditional knock-in of pathogenic mutations in Polr3a, which encodes the largest Pol III subunit, specifically in Olig2-expressing cells in the CNS, leads to reduce growth, neurobehavioral deficits and hypomyelination in multiple regions of the brain and spinal cord. Moreover, electron microscopy along with an analysis of oligodendrocyte cell populations and specific protein markers has identified two distinct mechanisms contributing to disease pathogenesis: Defective differentiation of oligodendrocyte precursor cells (OPCs) and defective function of mature oligodendrocytes (OLs). Nevertheless, the molecular mechanisms of pathogenesis remain poorly characterized. To advance understanding in this regard, the long- term goals of this work are to determine how the Pol III transcriptome has been altered in OPCs and OLs in the mutant mice and to examine the consequences of these changes for protein synthesis and for the proteome of OPCs and OLs and the lipidome of myelin. Additionally, we will test a model for genetic suppression of Pol III- related leukodystrophy phenotypes in the mice. Together, these studies will inform how decreasing Pol III transcription interferes with oligodendrogenesis and myelination, yielding important mechanistic insights, and a potential therapeutic approach that may benefit patients with Pol III-related leukodystrophy.