Autophagy inhibition promotes SNCA/alpha-synuclein release and transfer via extracellular vesicles with a hybrid autophagosome-exosome-like phenotype
Autophagy inhibition promotes SNCA/alpha-synuclein release and transfer via extracellular vesicles with a hybrid autophagosome-exosome-like phenotype
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DOI：
10.1080/15548627.2017.1395992
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发表时间：
2018-01-01
期刊：
影响因子：
13.3
通讯作者：
Klucken, Jochen
中科院分区：
文献类型：
--
作者：
Minakaki, Georgia;Menges, Stefanie;Klucken, Jochen
自噬-溶酶体途径（ALP）调节胞浆蛋白SNCA/ α -突触核蛋白的细胞内稳态，并在突触核蛋白病中受损，包括帕金森病和路易体痴呆（DLB）。新出现的证据表明ALP影响SNCA的释放，但潜在的细胞机制尚不清楚。一些研究在外泌体/细胞外囊泡（EV）中发现了SNCA。ev在多泡体室中产生，并在与质膜融合时释放，或通过碱性磷酸酶清除。因此，我们假设抑制ALP清除1)通过增加多泡体内容物的细胞外穿梭来增强细胞内SNCA的释放，2)改变细胞内生化特征，3)促进SNCA细胞间转移。事实上，ALP抑制增加了细胞外和细胞内SNCA的比例，并上调了神经元细胞中SNCA与ev的关联。超微结构分析显示广泛、融合的多泡体-自噬体室。生化鉴定显示存在自噬体相关蛋白，如LC3-II和SQSTM1。在人脑脊液（CSF） ev中也发现了这种独特的“自噬体-外泌体样”特征。将CSF衍生的含有SNCA的ev单次皮质内注射到小鼠体内后，人SNCA与核内体和神经元标记共定位。DLB患者脑脊液EV注射后，观察到明显的SNCA免疫反应性和较高数量的神经元SNCA包涵体。总之，本研究提供了令人信服的证据，a) ALP抑制增加了神经元ev中的SNCA， b) ev中存在不同的ALP成分，以及c) CSF ev在体内将SNCA从细胞转移到细胞。因此，巨噬/自噬可能调节EV蛋白组成，从而调节突触核蛋白病的进展。
The autophagy-lysosome pathway (ALP) regulates intracellular homeostasis of the cytosolic protein SNCA/alpha-synuclein and is impaired in synucleinopathies, including Parkinson disease and dementia with Lewy bodies (DLB). Emerging evidence suggests that ALP influences SNCA release, but the underlying cellular mechanisms are not well understood. Several studies identified SNCA in exosome/extracellular vesicle (EV) fractions. EVs are generated in the multivesicular body compartment and either released upon its fusion with the plasma membrane, or cleared via the ALP. We therefore hypothesized that inhibiting ALP clearance 1) enhances SNCA release via EVs by increasing extracellular shuttling of multivesicular body contents, 2) alters EV biochemical profile, and 3) promotes SNCA cell-to-cell transfer. Indeed, ALP inhibition increased the ratio of extra- to intracellular SNCA and upregulated SNCA association with EVs in neuronal cells. Ultrastructural analysis revealed a widespread, fused multivesicular body-autophagosome compartment. Biochemical characterization revealed the presence of autophagosome-related proteins, such as LC3-II and SQSTM1. This distinct "autophagosome-exosome-like" profile was also identified in human cerebrospinal fluid (CSF) EVs. After a single intracortical injection of SNCA-containing EVs derived from CSF into mice, human SNCA colocalized with endosome and neuronal markers. Prominent SNCA immunoreactivity and a higher number of neuronal SNCA inclusions were observed after DLB patient CSF EV injections. In summary, this study provides compelling evidence that a) ALP inhibition increases SNCA in neuronal EVs, b) distinct ALP components are present in EVs, and c) CSF EVs transfer SNCA from cell to cell in vivo. Thus, macroautophagy/auto phagy may regulate EV protein composition and consequently progression in synucleinopathies.