An aggregate-inducing peripherin isoform generated through intron retention is upregulated in amyotrophic lateral sclerosis and associated with disease pathology
An aggregate-inducing peripherin isoform generated through intron retention is upregulated in amyotrophic lateral sclerosis and associated with disease pathology
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DOI：
10.1523/jneurosci.3222-07.2008
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发表时间：
2008-02-20
影响因子：
5.3
通讯作者：
Robertson, Janice
中科院分区：
文献类型：
--
作者：
Xiao, Shangxi;Tjostheim, Sonja;Robertson, Janice
神经元中间丝蛋白外周蛋白是肌萎缩侧索硬化症 (ALS) 中泛素化包涵体和轴突球体的组成部分。外周蛋白的过度表达会导致转基因小鼠的运动神经元变性，并且在 ALS 病例中已发现外周蛋白基因内的变异。我们之前已经证明，在表达突变超氧化物歧化酶-1的转基因小鼠中，神经毒性外周蛋白剪接变体的异常表达。这些发现表明 ALS 中可能发生外周蛋白剪接异常。在当前的研究中，通过人神经元 RNA 的逆转录 PCR 鉴定了外周蛋白剪接变体，并使用蛋白质印迹分析和免疫细胞化学比较了对照和 ALS 脊髓之间的表达。使用这种方法，我们鉴定了一种保留内含子 3 和 4 的新型外周蛋白转录物，可产生 28 kDa 剪接亚型，命名为 Per 28。使用 Per 28 特异性抗体，我们表明该亚型以低化学计量水平从外周蛋白基因表达，然而，当其表达上调时会导致外周蛋白聚集。重要的是，我们发现，与对照组相比，ALS 中 Per 28 的表达在 mRNA 和蛋白质水平上都有上调，并且 Per 28 与疾病病理学相关，特别是圆形内含物。这些发现首次证实 ALS 中存在外周蛋白剪接异常，从而产生易于聚集的剪接亚型。
The neuronal intermediate filament protein peripherin is a component of ubiquitinated inclusions and of axonal spheroids in amyotrophic lateral sclerosis (ALS). Overexpression of peripherin causes motor neuron degeneration in transgenic mice and variations within the peripherin gene have been identified in ALS cases. We have shown previously the abnormal expression of a neurotoxic peripherin splice variant in transgenic mice expressing mutant superoxide dismutase-1. These findings indicated that abnormalities of peripherin splicing may occur in ALS. In the current study, peripherin splice variants were identified by reverse transcription-PCR of human neuronal RNA and comparisons in expression made between control and ALS spinal cord using Western blot analysis and immunocytochemistry. Using this approach we have identified a novel peripherin transcript retaining introns 3 and 4 that results in a 28 kDa splice isoform, designated Per 28. Using an antibody specific to Per 28, we show that this isoform is expressed at low stoichiometric levels from the peripherin gene, however causes peripherin aggregation when its expression is upregulated. Importantly we show an upregulation of Per 28 expression in ALS compared with controls, at both the mRNA and protein levels, and that Per 28 is associated with disease pathology, specifically round inclusions. These findings are the first to establish that peripherin splicing abnormalities occur in ALS, generating aggregation-prone splice isoforms.