Myosin VI isoform localized to clathrin-coated vesicles with a role in clathrin-mediated endocytosis

Myosin VI is involved in membrane traffic and dynamics and is the only myos in known to move towards the minus end of actin filaments. Splice variants of myosin VI with a large insert in the tail domain were specifically expre ssed in polarized cells containing microvilli. In these polarized cells, en dogenous myosin VI containing the large insert was concentrated at the apic al domain co-localizing with clathrin-coated pits/vesicles. Using full-leng th myosin VI and deletion mutants tagged with green fluorescent protein (GF P) we have shown that myosin VI associates and co-localizes with clathrin-c oated pits/vesicles by its C-terminal tail. Myosin VI, precipitated from wh ole cytosol, was present in a protein complex containing adaptor protein (A P)-2 and clathrin, and enriched in purified clathrin-coated vesicles. Over- expression of the tail domain of myosin VI containing the large insert in f ibroblasts reduced transferrin uptake in transiently and stably transfected cells by > 50%. Myosin VI is the firsts motor protein to be identified ass ociated with clathrin-coated pits/vesicles and shown to modulate clathrin-m ediated endocytosis.