The necdin interactome: evaluating the effects of amino acid substitutions and cell stress using proximity-dependent biotinylation (BioID) and mass spectrometry
The necdin interactome: evaluating the effects of amino acid substitutions and cell stress using proximity-dependent biotinylation (BioID) and mass spectrometry
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DOI：
10.1007/s00439-020-02193-9
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发表时间：
2020-06-11
期刊：
影响因子：
5.3
通讯作者：
Wevrick, Rachel
中科院分区：
文献类型：
--
作者：
Sanderson, Matthea R.;Badior, Katherine E.;Wevrick, Rachel
Prader-Willi综合征（PWS）是由染色体15q11 - 15q13上的一组印记基因的功能丧失引起的神经发育障碍。这些基因之一，NDN，编码necdin，一种对神经元分化和存活很重要的蛋白质。小鼠中Ndn的缺失导致神经系统的形成和功能缺陷。Necdin是黑色素瘤相关抗原基因（法师）蛋白家族的成员。法师蛋白的功能高度依赖于它们与其它蛋白的相互作用，特别是法师蛋白与E3泛素连接酶和去泛素化酶相互作用形成MAGE-RING E3连接酶-去泛素化酶复合物。在这里，我们使用邻近依赖性生物素鉴定（BioID）和质谱（MS）来确定necdin蛋白的蛋白质-蛋白质相互作用（相互作用组）网络。这一过程产生了新的以及已知的necdin近端蛋白，这些蛋白聚集成蛋白质网络。接下来，我们使用BioID-MS来定义携带编码变体的necdin蛋白的相互作用组。变异necdin蛋白的相互作用是从野生型necdin不同。BioID-MS不仅是鉴定蛋白质-蛋白质相互作用的有用工具，而且还可以分析未知意义的变体对遗传疾病相关蛋白质相互作用组的影响。
Prader-Willi syndrome (PWS) is a neurodevelopmental disorder caused by the loss of function of a set of imprinted genes on chromosome 15q11-15q13. One of these genes,NDN, encodes necdin, a protein that is important for neuronal differentiation and survival. Loss ofNdnin mice causes defects in the formation and function of the nervous system. Necdin is a member of the melanoma-associated antigen gene (MAGE) protein family. The functions of MAGE proteins depend highly on their interactions with other proteins, and in particular MAGE proteins interact with E3 ubiquitin ligases and deubiquitinases to form MAGE-RING E3 ligase-deubiquitinase complexes. Here, we used proximity-dependent biotin identification (BioID) and mass spectrometry (MS) to determine the network of protein-protein interactions (interactome) of the necdin protein. This process yielded novel as well as known necdin-proximate proteins that cluster into a protein network. Next, we used BioID-MS to define the interactomes of necdin proteins carrying coding variants. Variant necdin proteins had interactomes that were distinct from wildtype necdin. BioID-MS is not only a useful tool to identify protein-protein interactions, but also to analyze the effects of variants of unknown significance on the interactomes of proteins involved in genetic disease.