The absorption and endocytosis mechanism of flaxseed cyclic peptide CLQ is mediated by its binding to clathrin.

The absorption mechanism of cyclic peptides at the molecular level remains unclear. Previously, we found that flaxseed-derived cyclic peptide was absorbed into cells through endocytosis pathway dependent on clathrin. This study aimed to elucidate how CLQ triggers clathrin-mediated endocytosis. CLQ was labeled with FITC fluorescent probe, and results showed that CLQ was absorbed in a concentration- and time-dependent manner. Molecular docking, immunofluorescence colocalization and surface plasmon resonance showed that CLQ firmly binds to clathrin, even at the cellular level. The expression of key genes and proteins involved in clathrin-mediated endocytic vesicle formation was upregulated when cells absorbing CLQ, including CLTC, AP2M1, DNM2, and EPS15, whereas co-treatment with clathrin inhibitor (CPZ), this upregulation was suppressed. Above results suggested that CLQ binds with clathrin, then recruiting AP2M1, DNM2, and EPS15 to form endocytic vesicle, inducing the occurrence of clathrin-mediated endocytosis. These findings established a theoretical foundation for the development and utilization of dietary cyclic peptides.