MRC Toxicology Unit, University of Cambridge, Cambridge, UK.
Department of Biochemistry, Boston University, Boston, MA, USA.
Methods Mol Biol. 2023;2693:163-174. doi: 10.1007/978-1-0716-3342-7_13.
Molecular chaperones can interact with multiple proteins to form large networks. Understanding these interactions may shed light on the complexity of the chaperone functions. Here we developed a protocol for a modified proximity ligation-based methodology (PLA) for the detection of protein-protein interactions in order to understand how the Hsp70-Bag3 complex interacts with components of the Hippo signaling pathway. These experiments helped to elucidate the mechanisms of transmission of the proteotoxic stress signal to the Hippo pathway. The modified PLA technology has many advantages compared to co-immunoprecipitation protocols. It has higher sensitivity, is quantitative, and can be done in a 96-well format.
分子伴侣可以与多种蛋白质相互作用形成大型网络。了解这些相互作用可能有助于揭示伴侣功能的复杂性。在这里，我们开发了一种改良的基于邻近连接的方法 (PLA) 用于检测蛋白质-蛋白质相互作用，以了解 Hsp70-Bag3 复合物如何与 Hippo 信号通路的成分相互作用。这些实验有助于阐明将蛋白毒性应激信号传递到 Hippo 通路的机制。与共免疫沉淀方案相比，改良的 PLA 技术具有许多优势。它具有更高的灵敏度、定量性并且可以在 96 孔格式中进行。
