Department of Biochemistry and Molecular Biology, University of Melbourne, Parkville, Victoria 3010, Australia.
Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, Victoria 3010, Australia.
Sci Data. 2018 Nov 20;5:180262. doi: 10.1038/sdata.2018.262.
Ataxin-1 mutation, arising from a polyglutamine (polyQ) tract expansion, is the underlying genetic cause of the late-onset neurodegenerative disease Spinocerebellar ataxia type 1 (SCA1). To identify protein partners of polyQ-ataxin-1 in neuronal cells under control or stress conditions, here we report our complementary proteomics strategies of proximity-dependent biotin identification (BioID) and affinity purification (via GFP-Trap pulldown) in Neuro-2a cells expressing epitope-tagged forms of ataxin-1[85Q]. These approaches allowed our enrichment of proximal proteins and interacting partners, respectively, with the subsequent protein identification performed by liquid chromatography-MS/MS. Background proteins, not dependent on the presence of the polyQ-ataxin-1 protein, were additionally defined by their endogenous biotinylation (for the BioID protocol) or by their non-specific interaction with GFP only (in the GFP-Trap protocol). All datasets were generated from biological replicates. Following the removal of the identified background proteins from the acquired protein lists, our experimental design has captured a comprehensive polyQ-ataxin-1 proximal and direct protein partners under normal and stress conditions. Data are available via ProteomeXchange, with identifier PXD010352.
Ataxin-1 突变是一种多聚谷氨酰胺（polyQ）序列扩展，是迟发性神经退行性疾病脊髓小脑共济失调 1 型（SCA1）的潜在遗传原因。为了鉴定在对照或应激条件下神经元细胞中多聚谷氨酰胺-ataxin-1 的蛋白伴侣，我们在这里报告了我们在表达表位标记形式的 ataxin-1[85Q]的 Neuro-2a 细胞中进行的邻近依赖性生物素鉴定（BioID）和亲和纯化（通过 GFP-Trap 下拉）的互补蛋白质组学策略。这些方法分别允许我们富集近端蛋白和相互作用的伴侣，随后通过液相色谱-MS/MS 进行蛋白质鉴定。背景蛋白不依赖于多聚谷氨酰胺-ataxin-1 蛋白的存在，通过其内源生物素化（对于 BioID 方案）或仅通过其与 GFP 的非特异性相互作用（在 GFP-Trap 方案中）来定义。所有数据集均来自生物重复。从获得的蛋白质列表中去除鉴定的背景蛋白后，我们的实验设计捕获了正常和应激条件下多聚谷氨酰胺-ataxin-1 近端和直接蛋白伴侣的综合信息。数据可通过 ProteomeXchange 获得，标识符为 PXD010352。
