MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom.
PLoS Genet. 2009 Dec;5(12):e1000773. doi: 10.1371/journal.pgen.1000773. Epub 2009 Dec 18.
Spinal muscular atrophy is a severe motor neuron disease caused by inactivating mutations in the SMN1 gene leading to reduced levels of full-length functional SMN protein. SMN is a critical mediator of spliceosomal protein assembly, and complete loss or drastic reduction in protein leads to loss of cell viability. However, the reason for selective motor neuron degeneration when SMN is reduced to levels which are tolerated by all other cell types is not currently understood. Widespread splicing abnormalities have recently been reported at end-stage in a mouse model of SMA, leading to the proposition that disruption of efficient splicing is the primary mechanism of motor neuron death. However, it remains unclear whether splicing abnormalities are present during early stages of the disease, which would be a requirement for a direct role in disease pathogenesis. We performed exon-array analysis of RNA from SMN deficient mouse spinal cord at 3 time points, pre-symptomatic (P1), early symptomatic (P7), and late-symptomatic (P13). Compared to littermate control mice, SMA mice showed a time-dependent increase in the number of exons showing differential expression, with minimal differences between genotypes at P1 and P7, but substantial variation in late-symptomatic (P13) mice. Gene ontology analysis revealed differences in pathways associated with neuronal development as well as cellular injury. Validation of selected targets by RT-PCR confirmed the array findings and was in keeping with a shift between physiologically occurring mRNA isoforms. We conclude that the majority of splicing changes occur late in SMA and may represent a secondary effect of cell injury, though we cannot rule out significant early changes in a small number of transcripts crucial to motor neuron survival.
脊髓性肌萎缩症是一种严重的运动神经元疾病，由 SMN1 基因的失活突变引起，导致全长功能性 SMN 蛋白水平降低。SMN 是剪接体蛋白组装的关键介质，蛋白质完全缺失或大幅减少会导致细胞活力丧失。然而，当 SMN 减少到所有其他细胞类型都能耐受的水平时，运动神经元选择性退化的原因目前尚不清楚。最近在 SMA 的小鼠模型中晚期报道了广泛的剪接异常，导致人们提出有效剪接的破坏是运动神经元死亡的主要机制。然而，在疾病早期是否存在剪接异常尚不清楚，如果这是疾病发病机制中的直接作用，则需要存在剪接异常。我们对 3 个时间点（无症状前（P1）、早期症状（P7）和晚期症状（P13））的 SMN 缺陷型小鼠脊髓 RNA 进行了外显子组分析。与同窝对照小鼠相比，SMA 小鼠表现出随时间推移差异表达的外显子数量增加，P1 和 P7 之间基因型之间的差异最小，但晚期症状（P13）小鼠的差异较大。基因本体论分析显示与神经元发育以及细胞损伤相关的途径存在差异。通过 RT-PCR 对选定的靶标进行验证，证实了阵列发现，并与生理发生的 mRNA 同工型之间的转变一致。我们得出结论，大多数剪接变化发生在 SMA 晚期，可能是细胞损伤的继发效应，尽管我们不能排除对运动神经元存活至关重要的少数转录本存在早期显著变化。
