Neuronal subclass-selective proteomic analysis in Caenorhabditis elegans
Neuronal subclass-selective proteomic analysis in Caenorhabditis elegans
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DOI：
10.1038/s41598-020-70692-w
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发表时间：
2020-08-13
影响因子：
4.6
通讯作者：
Ueda, Mitsuyoshi
中科院分区：
文献类型：
--
作者：
Aburaya, Shunsuke;Yamauchi, Yuji;Ueda, Mitsuyoshi
神经元分为许多子类，每个子类表现出不同的形态、表达模式、连接性和功能。蛋白质合成的变化对于神经元功能至关重要。因此，分析单个神经元亚类中的蛋白质表达模式将阐明记忆和其他功能的分子机制。在这项研究中，我们使用神经元亚类选择性蛋白质组分析和细胞选择性生物正交非规范氨基酸标记。我们选择秀丽隐杆线虫作为模式生物，因为它显示出多样化的神经元功能和简单的神经回路。我们对秀丽隐杆线虫中调节趋热性的所有神经元或 AFD 亚类神经元进行了蛋白质组学分析。突变型苯丙氨酰 tRNA 合成酶 (MuPheRS) 在所有神经元或 AFD 亚类神经元中选择性表达，叠氮基苯丙氨酸被整合到感兴趣细胞的蛋白质中。富集叠氮化物标记的蛋白质并进行蛋白质组分析。我们分别从所有神经元和 AFD 亚类神经元中产生 MuPheRS 的菌株中鉴定出 4,412 和 1,834 个蛋白质。 F23B2.10（含环型结构域的蛋白质）仅在神经元细胞富集的蛋白质组分析中被鉴定。我们在F23B2.10 5'调控区的控制下表达GFP，并发现GFP在神经元中表达。我们期望更多的单神经元特异性蛋白质组数据将阐明蛋白质组成和丰度如何影响神经元亚类的特征。
Neurons are categorised into many subclasses, and each subclass displays different morphology, expression patterns, connectivity and function. Changes in protein synthesis are critical for neuronal function. Therefore, analysing protein expression patterns in individual neuronal subclass will elucidate molecular mechanisms for memory and other functions. In this study, we used neuronal subclass-selective proteomic analysis with cell-selective bio-orthogonal non-canonical amino acid tagging. We selected Caenorhabditis elegans as a model organism because it shows diverse neuronal functions and simple neural circuitry. We performed proteomic analysis of all neurons or AFD subclass neurons that regulate thermotaxis in C. elegans. Mutant phenylalanyl tRNA synthetase (MuPheRS) was selectively expressed in all neurons or AFD subclass neurons, and azido-phenylalanine was incorporated into proteins in cells of interest. Azide-labelled proteins were enriched and proteomic analysis was performed. We identified 4,412 and 1,834 proteins from strains producing MuPheRS in all neurons and AFD subclass neurons, respectively. F23B2.10 (RING-type domain-containing protein) was identified only in neuronal cell-enriched proteomic analysis. We expressed GFP under the control of the 5 ' regulatory region of F23B2.10 and found GFP expression in neurons. We expect that more single-neuron specific proteomic data will clarify how protein composition and abundance affect characteristics of neuronal subclasses.