Moesin Regulates the Trafficking of Nascent Clathrin-coated Vesicles.

Abstract

Clathrin-coated vesicles are responsible for the trafficking of several internalized biological cargos. We have observed that the endogenous F-actin-linker moesin co-distributes with constitutive components of clathrin-coated structures.Totalinternal reflection fluorescence microscopy studies have shown that short interference RNA of moesin enhances the lateral movement of clathrincoated structures and provokes their abnormal clustering. The aggregation of clathrin-coated structures has also been observedin cells overexpressing N-moesin, a dominant-negative construct unable to bind to F-actin. Only overexpressed moesin constructs with an intact phosphatidylinositol 4,5-bisphosphate-binding domain co-distribute with clathrin-coated structures. Hence, this N-terminal domain is mostly responsible for moesin/clathrincoated structure association. Biochemical endosome fractioning together with total internal reflection fluorescence microscopy comparative studies, between intact cells and plasma-membrane sheets, indicate that moesin knockdown provokes the accumulation of endocytic rab5-clathrin-coated vesicles carrying the transferrin receptor. The altered trafficking of these endocytic rab5- clathrin-coated vesicles accounts for a transferrin receptor recycling defect that reduces cell-surface expression of the transferrin receptorandincreases theamount of sequestered transferrin ligand. Therefore, we propose that moesin is a clathrin-coated vesicle linker that drives cargo trafficking and acts on nascent rab5- clathrin-coated vesicles by simultaneously binding to clathrincoated vesicle-associated phosphatidylinositol 4,5-bisphosphate and actin cytoskeleton. Hence, functional alterations of moesin may be involved in pathological disorders associated with clathrinmediated internalization or receptor recycling.