PDF(Pubmed)
Abstract：
Contact sites between lipid droplets and other organelles are essential for cellular lipid and energy homeostasis upon metabolic demands. Detection of these contact sites at the nanometer scale over time in living cells is challenging. We developed a tool kit for detecting contact sites based on fluorogen-activated bimolecular complementation at CONtact sites, FABCON, using a reversible, low-affinity split fluorescent protein, splitFAST. FABCON labels contact sites with minimal perturbation to organelle interaction. Via FABCON, we quantitatively demonstrated that endoplasmic reticulum (ER)- and mitochondria (mito)-lipid droplet contact sites are dynamic foci in distinct metabolic conditions, such as during lipid droplet biogenesis and consumption. An automated analysis pipeline further classified individual contact sites into distinct subgroups based on size, likely reflecting differential regulation and function. Moreover, FABCON is generalizable to visualize a repertoire of organelle contact sites including ER-mito. Altogether, FABCON reveals insights into the dynamic regulation of lipid droplet-organelle contact sites and generates new hypotheses for further mechanistical interrogation during metabolic regulation.
摘要：
脂滴和其他细胞器之间的接触位点对于代谢需求时的细胞脂质和能量稳态至关重要。在活细胞中随时间在纳米尺度上检测这些接触位点是具有挑战性的。我们开发了一个工具包，用于检测基于荧光激活的双分子互补的接触位点在CONtact位点，FABCON,使用可逆的，低亲和力分裂荧光蛋白，splitFAST。FABCON标记接触位点，对细胞器相互作用的扰动最小。通过FABCON,我们定量地证明了内质网（ER）-和线粒体（mito）-脂滴接触位点是不同代谢条件下的动态病灶，例如在脂滴生物生成和消耗期间。自动分析管道根据大小进一步将各个接触点分为不同的子组，可能反映了不同的调节和功能。此外，FABCON可概括为可视化包括ER-mito的细胞器接触位点。总之,FABCON揭示了对脂滴-细胞器接触位点的动态调节的见解，并为代谢调节过程中的进一步机械询问提供了新的假设。
