Uptake of CD36 and its natural and therapeutic ligands by clathrin-mediated endocytosis

Abstract

The scavenger receptor CD36 mediates cellular uptake of diverse extracellular ligands including long-chain fatty acids. Recent work has demonstrated that CD36 is also required for uptake of bulky, amphipathic therapeutic compounds including proteolysis-targeting chimeras (PROTACs). However, because the mechanism by which cells internalize CD36 itself remains unknown, how its diverse ligands enter cells has remained mysterious. CD36 has been thought to associate with caveolae, flask-like invaginations of the plasma membrane, yet recent reports have cast doubt on the role of these structures in protein trafficking. Using live cell imaging, we show that CD36 strongly colocalizes with sites of clathrin-mediated endocytosis, where it concentrates both long-chain fatty acids and PROTACs. In contrast, neither CD36 nor its ligands colocalized substantially with caveolae. Further, we identify previously unrecognized binding interactions between CD36 and the clathrin machinery, which help to explain recruitment of CD36 and its ligands to endocytic sites. Importantly, disruption of clathrin assembly inhibits endocytic uptake of both long-chain fatty acids and PROTACs, and abolished PROTAC-induced degradation of multiple target proteins that utilize diverse E3 enzymes. These results suggest that clathrin-mediated endocytosis is essential for internalization of CD36 and its ligands, offering new insights into diverse diseases that rely on CD36 for ligand import, and potentially inspiring new strategies for optimizing targeted protein degraders.