The Cdc42 target ACK2 directly interacts with clathrin and influences clathrin assembly.

The Ras-related GTP-binding protein Cdc42 has been implicated in a diversity of biological functions including the regulation of intracellular trafficking and endocytosis. While screening for Cdc42 targets that influence these activities, we identified the protein-tyrosine kinase ACK2 (for activated Cdc42-associated kinase 2) as a new binding partner for clathrin. ACK2 binds clathrin via a domain that is conserved among a number of other clathrin-binding proteins including the arrestins and AP-2. Overexpression of ACK2 in NIH3T3 cells results in an inhibition of transferrin receptor endocytosis because of a competition between ACK2 and AP-2 for clathrin. Activated Cdc42 weakens the interaction between ACK2 and clathrin and thus reverses the ACK2-mediated inhibition of endocytosis. Overexpression of ACK2 increases the amount of clathrin present in fractions enriched in clathrin-coated vesicles. Taken together, our data suggest that ACK2 may represent a novel clathrin-assembly protein and participate in the regulation of receptor-mediated endocytosis.