Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia.
QIMR Berghofer Medical Research Institute, Herston, Australia.
Elife. 2021 Feb 16;10:e64631. doi: 10.7554/eLife.64631.
Protein interaction networks are crucial for complex cellular processes. However, the elucidation of protein interactions occurring within highly specialised cells and tissues is challenging. Here, we describe the development, and application, of a new method for proximity-dependent biotin labelling in whole zebrafish. Using a conditionally stabilised GFP-binding nanobody to target a biotin ligase to GFP-labelled proteins of interest, we show tissue-specific proteomic profiling using existing GFP-tagged transgenic zebrafish lines. We demonstrate the applicability of this approach, termed BLITZ (Biotin Labelling In Tagged Zebrafish), in diverse cell types such as neurons and vascular endothelial cells. We applied this methodology to identify interactors of caveolar coat protein, cavins, in skeletal muscle. Using this system, we defined specific interaction networks within in vivo muscle cells for the closely related but functionally distinct Cavin4 and Cavin1 proteins.
蛋白质相互作用网络对于复杂的细胞过程至关重要。然而，阐明发生在高度特化的细胞和组织中的蛋白质相互作用具有挑战性。在这里，我们描述了一种新的方法的开发和应用，用于在整个斑马鱼中进行邻近依赖性生物素标记。使用条件稳定的 GFP 结合纳米体将生物素连接酶靶向 GFP 标记的感兴趣的蛋白质，我们展示了使用现有的 GFP 标记的转基因斑马鱼系进行组织特异性蛋白质组学分析。我们证明了这种方法（称为 BLITZ（生物素标记在标记的斑马鱼中））在神经元和血管内皮细胞等多种细胞类型中的适用性。我们将这种方法应用于鉴定骨骼肌中腔囊泡外衣蛋白 cavin 的相互作用蛋白。使用该系统，我们为密切相关但功能不同的 Cavin4 和 Cavin1 蛋白在体内肌肉细胞内定义了特定的相互作用网络。
Zotero 插件
