Cathepsin D deficiency induces cytoskeletal changes and affects cell migration pathways in the brain
Cathepsin D deficiency induces cytoskeletal changes and affects cell migration pathways in the brain
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DOI：
10.1016/j.nbd.2012.10.004
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发表时间：
2013-02-01
影响因子：
6.1
通讯作者：
Tyynela，Jaana
中科院分区：
文献类型：
--
作者：
Koch，Sabine;Scifo，Enzo;Tyynela，Jaana
组织蛋白酶D缺乏症是一种以神经元和髓鞘极度丧失为特征的致命性神经退行性疾病。我们以前的研究已经证明，突触的结构和功能变化是疾病发病机制的核心。因此，我们采取了一种系统的方法来检测组织蛋白D基因敲除小鼠的突触蛋白质组，其中突触的病理类似于人类患者。我们对24日龄组织蛋白酶D基因敲除小鼠和对照小鼠的突触体组分进行了定量质谱分析。在大约600个已鉴定的蛋白质中，43个在组织蛋白酶D基因敲除小鼠中以不同的数量存在(P<；0.05，以三个生物重复测量)。我们使用蛋白质相互作用数据连接和桥接这43个蛋白质，并用大脑特定的基因表达信息覆盖网络。随后，我们将网络与基因本体论、途径、表型和疾病参与叠加在一起，从而构建了一个以疾病蛋白为中心的动态网络，并预测了功能模块。定量Western blotting或定性免疫组织化学证实了蛋白质水平的测量变化，以及一些生物信息预测的变化。这一联合方法显示了以前与疾病无关的不同细胞实体的变化，包括微管相关的细胞骨架和细胞投射组织。细胞扩散和伤口愈合分析证实，在组织蛋白酶D缺乏的细胞中，与局部粘连和整合素组装的分布变化有关的空间取向严重受损。这些变化可能与组织蛋白酶D缺乏所观察到的突触改变和神经元变性有关。
Cathepsin D deficiency is a fatal neurodegenerative disease characterized by extreme loss of neurons and myelin. Our previous studies have demonstrated that structural and functional alterations in synapses are central to the disease pathogenesis. Therefore, we took a systematic approach to examine the synaptic proteome in cathepsin D knock-out mice, where the synaptic pathology resembles that of human patients. We applied quantitative mass spectrometry analysis on synaptosomal fractions isolated from cathepsin D knock-out and control mice at the age of 24days. From the approximately 600 identified proteins, 43 were present in different amounts (P<0.05, measured in triple biological replicates) in cathepsin D knock-out mice compared to controls. We connected and bridged these 43 proteins using protein interaction data, and overlaid the network with brain specific gene expression information. Subsequently, we superimposed the network with Gene Ontology, pathway, phenotype and disease involvement, allowing construction of a dynamic, disease-protein centered network and prediction of functional modules. The measured changes in the protein levels, as well as some of the bioinformatically predicted ones, were confirmed by quantitative Western blotting or qualitative immunohistochemistry. This combined approach indicated alterations in distinct cellular entities, previously not associated with the disease, and including microtubule associated cytoskeleton and cell projection organization. Cell spreading and wound healing assays confirmed strongly compromised spatial orientation, associated with changes in distribution of focal adhesions and integrin assembly, in cathepsin D deficient cells. These changes might contribute to commencement of synaptic alterations and neuronal degeneration observed in cathepsin D deficiency.