Institute for Genetics, University of Cologne, Cologne, Germany.
Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany.
Methods Mol Biol. 2022;2525:197-205. doi: 10.1007/978-1-0716-2473-9_14.
The contacts between the endoplasmic reticulum (ER) and mitochondria play a fundamental role in a wide variety of cellular processes, like the exchange of calcium and lipids between both organelles, as well as in apoptosis and in autophagy signaling. Despite their importance, due to their dynamic and heterogeneous nature, we still lack understanding of the molecular composition, structure, and regulation of these structures. In this chapter, we introduce a new bioluminescence resonance energy transfer (BRET)-based biosensor for the quantitative analysis of mitochondria-ER interorganellar distances without perturbing their natural environment, which we call MERLIN (mitochondria ER length indicator nanosensor). Here, we describe the rationale behind the MERLIN biosensor, detail the experimental setup and methodology, and provide tips for troubleshooting.
内质网 (ER) 和线粒体之间的接触在广泛的细胞过程中起着至关重要的作用，例如细胞器之间钙和脂质的交换，以及细胞凋亡和自噬信号转导。尽管它们很重要，但由于它们的动态和异质性，我们仍然缺乏对这些结构的分子组成、结构和调节的理解。在本章中，我们介绍了一种新的基于生物发光共振能量转移 (BRET) 的生物传感器，用于定量分析线粒体-ER 细胞器间距离，而不会干扰其自然环境，我们称之为 MERLIN（线粒体 ER 长度指示纳米传感器）。在这里，我们描述了 MERLIN 生物传感器的基本原理，详细说明了实验设置和方法，并提供了故障排除技巧。
