a Department of Neurology , Columbia University Medical Center , New York , NY , USA.
b Department of Neurology and Institute for Cell Engineering , Johns Hopkins University , Baltimore , MD , USA.
Autophagy. 2019 Jan;15(1):113-130. doi: 10.1080/15548627.2018.1509818. Epub 2018 Oct 12.
Heterozygous mutations in GBA, the gene encoding the lysosomal enzyme glucosylceramidase beta/β-glucocerebrosidase, comprise the most common genetic risk factor for Parkinson disease (PD), but the mechanisms underlying this association remain unclear. Here, we show that in Gba knockin mice, the L444P heterozygous Gba mutation triggers mitochondrial dysfunction by inhibiting autophagy and mitochondrial priming, two steps critical for the selective removal of dysfunctional mitochondria by autophagy, a process known as mitophagy. In SHSY-5Y neuroblastoma cells, the overexpression of L444P GBA impeded mitochondrial priming and autophagy induction when endogenous lysosomal GBA activity remained intact. By contrast, genetic depletion of GBA inhibited lysosomal clearance of autophagic cargo. The link between heterozygous GBA mutations and impaired mitophagy was corroborated in postmortem brain tissue from PD patients carrying heterozygous GBA mutations, where we found increased mitochondrial content, mitochondria oxidative stress and impaired autophagy. Our findings thus suggest a mechanistic basis for mitochondrial dysfunction associated with GBA heterozygous mutations. Abbreviations: AMBRA1: autophagy/beclin 1 regulator 1; BECN1: beclin 1, autophagy related; BNIP3L/Nix: BCL2/adenovirus E1B interacting protein 3-like; CCCP: carbonyl cyanide 3-chloroyphenylhydrazone; CYCS: cytochrome c, somatic; DNM1L/DRP1: dynamin 1-like; ER: endoplasmic reticulum; GBA: glucosylceramidase beta; GBA-PD: Parkinson disease with heterozygous GBA mutations; GD: Gaucher disease; GFP: green fluorescent protein; LC3B: microtubule-associated protein 1 light chain 3 beta; LC3B-II: lipidated form of microtubule-associated protein 1 light chain 3 beta; MitoGreen: MitoTracker Green; MitoRed: MitoTracker Red; MMP: mitochondrial membrane potential; MTOR: mechanistic target of rapamycin kinase; MYC: MYC proto-oncogene, bHLH transcription factor; NBR1: NBR1, autophagy cargo receptor; Non-GBA-PD: Parkinson disease without GBA mutations; PD: Parkinson disease; PINK1: PTEN induced putative kinase 1; PRKN/PARK2: parkin RBR E3 ubiquitin protein ligase; RFP: red fluorescent protein; ROS: reactive oxygen species; SNCA: synuclein alpha; SQSTM1/p62: sequestosome 1; TIMM23: translocase of inner mitochondrial membrane 23; TOMM20: translocase of outer mitochondrial membrane 20; VDAC1/Porin: voltage dependent anion channel 1; WT: wild type.
GBA 基因编码溶酶体酶β-葡萄糖脑苷脂酶，其杂合突变是帕金森病（PD）最常见的遗传风险因素，但这种关联的机制仍不清楚。在这里，我们表明，在 Gba 基因敲入小鼠中，L444P 杂合 Gba 突变通过抑制自噬和线粒体启动来触发线粒体功能障碍，这两个步骤对于通过自噬选择性去除功能失调的线粒体（称为线粒体自噬）至关重要。在 SHSY-5Y 神经母细胞瘤细胞中，当内源性溶酶体 GBA 活性保持完整时，L444P GBA 的过表达会阻碍线粒体启动和自噬诱导。相比之下，GBA 的基因缺失会抑制自噬小体的溶酶体清除。在携带杂合 GBA 突变的 PD 患者的死后脑组织中，证实了杂合 GBA 突变与受损的线粒体自噬之间的联系，我们发现线粒体含量增加、线粒体氧化应激和自噬受损。因此，我们的研究结果为 GBA 杂合突变相关的线粒体功能障碍提供了一种机制基础。缩写：AMBRA1：自噬/贝林 1 调节剂 1；BECN1：自噬相关的 beclin 1；BNIP3L/Nix：BCL2/腺病毒 E1B 相互作用蛋白 3 样；CCCP：羰基氰化物 3-氯苯腙；CYCS：细胞色素 c，体细胞；DNM1L/DRP1：dynamin 1 样；ER：内质网；GBA：β-葡萄糖脑苷脂酶；GBA-PD：携带杂合 GBA 突变的帕金森病；GD：戈谢病；GFP：绿色荧光蛋白；LC3B：微管相关蛋白 1 轻链 3β；LC3B-II：微管相关蛋白 1 轻链 3β 的脂化形式；MitoGreen：MitoTracker Green；MitoRed：MitoTracker Red；MMP：线粒体膜电位；MTOR：雷帕霉素靶蛋白激酶；MYC：原癌基因 MYC，bHLH 转录因子；NBR1：NBR1，自噬货物受体；Non-GBA-PD：无 GBA 突变的帕金森病；PD：帕金森病；PINK1：PTEN 诱导的假定激酶 1；PRKN/PARK2：parkin RBR E3 泛素蛋白连接酶；RFP：红色荧光蛋白；ROS：活性氧；SNCA：突触核蛋白 alpha；SQSTM1/p62：自噬体 1；TIMM23：线粒体内膜 23 转位酶；TOMM20：外线粒体膜 20 转位酶；VDAC1/ Porin：电压依赖性阴离子通道 1；WT：野生型。
