A clickable CoQ imaging probe reveals that cellular uptake and lysosomal trafficking depend on CD36 and NPC1.

Coenzyme Q (CoQ) is a crucial lipid-soluble antioxidant and electron transporter vital for mitochondrial respiration and cellular redox balance. Despite the role of CoQ in oxidative phosphorylation being well established, the mechanisms by which CoQ is internalized, distributed among subcellular compartments, and trafficked to mitochondria remain poorly defined. Here, we present the development of a minimally modified, azide-tagged CoQ analogue that enables high-resolution visualization of CoQ localization using fluorescence-based imaging. Using this probe, we focus our investigation on brown adipose tissue (BAT), a mitochondria-rich, highly metabolically active tissue with elevated CoQ demand. On a cellular level, we demonstrate that CoQ is internalized via receptor-mediated endocytosis, predominantly localizing to lysosomes. Genetic knockdown and pharmacological studies identify CD36 and NPC1 as essential transporters in this process. Our work provides both a technical advance for the redox biology field, with the development and characterization of a CoQ probe, and the essential new biological insight that NPC1 is linked to CoQ homeostasis and thus provides a foundation for further dissection of CoQ biology in health and disease.