The mechanism of cell damage by mutant superoxide dismutase 1 of human amyotrophic lateral sclerosis

我们将野生型和G37 R、G85 R、G93 A、S134 N四种突变型超氧化物歧化酶1（SOD 1）基因亚克隆到pcDNA 3和pEF-BOS两种表达载体上。将SOD 1基因转染COS 7细胞。转染后48 h，在转染有高启动子活性的pEF-BOS中的任何突变SOD 1基因的COS 7细胞的胞体中观察到突变SOD 1蛋白的聚集体。这些聚集体定位于内质网（ER）。聚集体的形成与这些细胞中线粒体和微管蛋白的异常分布有关。亚克隆到启动子活性较低的pcDNA 3上的突变SOD 1基因在COS 7细胞中不产生任何聚集体。然而，给药的lactacystin，蛋白酶体抑制剂，培养基诱导的突变体蛋白质的聚集在细胞转染突变体SOD 1基因的pcDNA 3。在转染野生型SOD 1基因的细胞中，未观察到聚集现象，提示突变型SOD 1蛋白的过度表达超过了蛋白酶体的降解能力，导致突变型蛋白在内质网中聚集。这些聚集体可诱导细胞内细胞器如线粒体的功能障碍。

We subcloned wild and four mutant (G37R, G85R, G93A, S134N) superoxide dismutase 1 (SOD1) genes to two expression vectors, pcDNA3 and pEF-BOS. Each SOD1 gene was transfected to COS 7 cells. Forty eight hours after transfection, aggregates of the mutant SOD1 proteins were observed in the perikarya of COS7 cells transfected with any mutant SOD1 gene in pEF-BOS, which has a high promoter activity. These aggregates were localized to endoplasmic reticulum (ER). The aggregate formation was associated with abnormal distribution of both mitochondria and tubulin in these cells. The mutant SOD1 genes subcloned to pcDNA3, which has a relatively low promotor activity, did not produce any aggregates in COS 7 cells. However, the administration of lactacystin, a proteasome inhibitor, to culture medium induced the aggregation of the mutant proteins in cells transfected with the mutant SOD1 genes in pcDNA3. No aggregation was observed in cells transfected with wild SOD1 gene in pcDNA3 or pEF-BOS.In conclusion, it is suggested that the over-expression of mutant SOD1 proteins beyond the capacity of degradation by proteasome results in the aggregation of the mutant proteins in ER. These aggregates may induce a dysfunction of intracellular organelles, such as mitochondria.