A FRET Autophagy Imaging Platform by Macrocyclic Amphiphile

Autophagy is a ubiquitous process of organelle interaction in eukaryotic cells, in which various organelles or proteins are recycled and operated through the autophagy pathway to ensure nutrient and energy homeostasis. Although numerous fluorescent probes have been developed to image autophagy, these environment-responsive probes suffer from inherent deficiencies such as inaccuracy and limited versatility. Here, we present a modular macrocyclic amphiphile Förster Resonance Energy Transfer (FRET) platform (SC6A12C/NCM, SN), constructed through the amphiphilic assembly of sulfonatocalix[6]arene (SC6A12C) with N-cetylmorpholine (NCM) for lysosome targeting. The hydrophobic fluorophore BPEA (FRET donor) was entrapped within the inner hydrophobic phase and showed strong fluorescence emission. Attributed to the broad-spectrum encapsulation of SC6A12C, three commercially available organelle probes (Mito-Tracker Red, ER Tracker Red, and RhoNox-1) were selected as SC6A12C guests (FRET acceptors). During autophagy process, the formation of intracellular host-guest complexes leads to strong FRET signal, allowing us to visualize the fusion of mitochondria, endoplasmic reticulum, and Golgi apparatus with lysosomes, respectively. This study provides a versatile and accessible platform for imaging organelle autophagy.