Glucocerebrosidase deficiency promotes protein aggregation through dysregulation of extracellular vesicles.
Glucocerebrosidase deficiency promotes protein aggregation through dysregulation of extracellular vesicles.
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DOI：
10.1371/journal.pgen.1007694
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发表时间：
2018-09
期刊：
影响因子：
4.5
通讯作者：
Pallanck LJ
中科院分区：
文献类型：
--
作者：
Thomas RE;Vincow ES;Merrihew GE;MacCoss MJ;Davis MY;Pallanck LJ
糖基神经酰胺酶β(GBA)基因突变与以蛋白质聚集为特征的神经退行性疾病密切相关。GBA编码溶酶体酶葡萄糖脑苷酶，该酶能分解葡萄糖神经酰胺。对GBA突变和蛋白质聚集之间的联系的一个常见解释是，葡萄糖神经酰胺的溶酶体积累导致自噬功能受损。我们通过测量Gba同源基因Gba1b缺失的果蝇突变体的蛋白质周转率和丰度，直接检验了这一假设。蛋白质组学分析显示，已知的自噬底物严重损害了自噬缺陷ATG7突变体的周转，但Gba1b突变体的周转几乎或没有总体减慢或丰度增加。同样，Gba1b突变体在线粒体蛋白周转方面没有丝裂原缺陷Parkin突变体所见的显著损害。在Gba1b突变体中，蛋白酶体活性、微自噬和内吞降解也没有受到影响。然而，我们发现与细胞外小泡(EVS)相关的蛋白质的周转率和丰度发生了惊人的变化，EV被认为是神经退行性疾病中蛋白质聚集体扩散的载体。这些变化是Gba1b突变所特有的，并不代表正常衰老的加速。对分离的EV进行Western blotting，证实了Gba1b突变体中EV蛋白的丰度增加，纳米颗粒跟踪分析表明，Gba1b突变体的EV数量是对照的6倍。Gba1b突变体中EV产生的遗传扰动抑制了蛋白质聚集，表明EV丰度的增加有助于蛋白质聚集的积累。总之，我们的发现表明，葡萄糖脑苷酶缺乏导致EV代谢的病理性变化，并可能促进蛋白质聚集体通过细胞外小泡传播。编码葡萄糖脑苷酶的GBA基因突变很常见，会增加患帕金森病的风险。一种被广泛接受的风险增加的解释是，通常由葡萄糖脑苷酶分解的脂肪物质会在细胞的回收中心--溶酶体中积聚，直到细胞无法再清除受损部分。在这一点上，应该在溶酶体中被破坏的蛋白质在整个细胞中形成大团块(聚集体)。我们使用没有葡萄糖脑苷酶的突变果蝇来测试这一理论，令我们惊讶的是，我们没有看到溶酶体失败的证据。在突变果蝇中，通常由溶酶体回收的蛋白质的破坏速度并没有减慢。相反，我们看到的证据表明，突变体的细胞可能产生了太多的细胞外小泡，即在细胞之间运输货物和信息的微小球体。一些研究人员还提出，随着帕金森病的恶化，细胞外小泡携带着在细胞间传播的蛋白质聚集体。我们的研究支持这一观点。这表明，聚集体通过细胞外小泡传播的增加，而不是溶酶体的失败，可能解释了为什么GBA突变增加了神经退行性疾病的风险。
Mutations in the glucosylceramidase beta (GBA) gene are strongly associated with neurodegenerative diseases marked by protein aggregation. GBA encodes the lysosomal enzyme glucocerebrosidase, which breaks down glucosylceramide. A common explanation for the link between GBA mutations and protein aggregation is that lysosomal accumulation of glucosylceramide causes impaired autophagy. We tested this hypothesis directly by measuring protein turnover and abundance in Drosophila mutants with deletions in the GBA ortholog Gba1b. Proteomic analyses revealed that known autophagy substrates, which had severely impaired turnover in autophagy-deficient Atg7 mutants, showed little to no overall slowing of turnover or increase in abundance in Gba1b mutants. Likewise, Gba1b mutants did not have the marked impairment of mitochondrial protein turnover seen in mitophagy-deficient parkin mutants. Proteasome activity, microautophagy, and endocytic degradation also appeared unaffected in Gba1b mutants. However, we found striking changes in the turnover and abundance of proteins associated with extracellular vesicles (EVs), which have been proposed as vehicles for the spread of protein aggregates in neurodegenerative disease. These changes were specific to Gba1b mutants and did not represent an acceleration of normal aging. Western blotting of isolated EVs confirmed the increased abundance of EV proteins in Gba1b mutants, and nanoparticle tracking analysis revealed that Gba1b mutants had six times as many EVs as controls. Genetic perturbations of EV production in Gba1b mutants suppressed protein aggregation, demonstrating that the increase in EV abundance contributed to the accumulation of protein aggregates. Together, our findings indicate that glucocerebrosidase deficiency causes pathogenic changes in EV metabolism and may promote the spread of protein aggregates through extracellular vesicles. Mutations in the GBA gene, which encodes the enzyme glucocerebrosidase, are common and increase the risk of Parkinson disease. A widely accepted explanation for the increased risk is that the fatty substance normally broken down by glucocerebrosidase builds up in the lysosome, which is the cell’s recycling center, until the cell can no longer get rid of damaged parts. At that point, proteins that should be destroyed in the lysosome form large clumps (aggregates) throughout the cell. We used mutant fruit flies without glucocerebrosidase to test this theory, and we were surprised to see no evidence that the lysosome was failing. The destruction of proteins usually recycled by the lysosome was not slowed down in the mutant flies. Instead, we saw evidence that the mutants’ cells might be producing too many extracellular vesicles, tiny spheres that transport cargo and messages from cell to cell. Some researchers have also suggested that extracellular vesicles carry the protein aggregates that spread between cells as Parkinson disease get worse. Our study supports this idea. It suggests that increased spread of aggregates through extracellular vesicles, rather than failure of the lysosome, might explain why GBA mutations increase the risk of neurodegenerative disease.
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DOI：
10.1111/tra.12016
发表时间：
2013-01
期刊：
Traffic (Copenhagen, Denmark)
影响因子：
--
作者：
Beckett K;Monier S;Palmer L;Alexandre C;Green H;Bonneil E;Raposo G;Thibault P;Le Borgne R;Vincent JP
通讯作者：
Vincent JP
影响因子：
7.8
作者：
Aflaki E;Moaven N;Borger DK;Lopez G;Westbroek W;Chae JJ;Marugan J;Patnaik S;Maniwang E;Gonzalez AN;Sidransky E
通讯作者：
Sidransky E
影响因子：
3.5
作者：
Awad, Ola;Sarkar, Chinmoy;Feldman, Ricardo A.
通讯作者：
Feldman, Ricardo A.
影响因子：
--
作者：
Alvarez-Erviti, Lydia;Rodriguez-Oroz, Maria C.;Schapira, Anthony H. V.
通讯作者：
Schapira, Anthony H. V.
影响因子：
14.9
作者：
UniProt Consortium
通讯作者：
UniProt Consortium