title: Cargo-binding makes a wild-type single-headed myosin-VI move processively.
abstract: Class VI myosin is an intracellular vesicle and organelle transporter that moves along actin filaments in a direction opposite to most other known myosin classes. The myosin-VI was expected to form a dimer to move processively along actin filaments with a hand-over-hand mechanism like other myosin organelle transporters. Recently, however, wild-type myosin-VI was demonstrated to be monomer and single-headed, casting a doubt on its processivity. By using single molecule techniques, we show that green-fluorescent-protein-tagged single-headed, wild-type myosin-VI does not move processively. However, when coupled to 200-nm polystyrene beads (comparable to intracellular vesicles in size) at a ratio of one head per bead, single-headed myosin-VI moves processively with large (40-nm) steps. The characteristics of this monomer-driven movement were different to that of artificial dimer-driven movement: Compared to the artificial dimer, the monomer-bead complex had a reduced stall force (1 pN compared to 2 pN), an average run length 2.5-fold shorter (91 nm compared to 220 nm) and load-dependent step size. Furthermore, we found that a monomer-bead complex moved more processively in a high viscous solution (40-fold higher than water) similar to cellular environment. Because the diffusion constant of the bead is 60-fold lower than myosin-VI heads alone in water, we propose a model in which the bead acts as a diffusional anchor for the myosin-VI, enhancing its rebinding following detachment and supporting processive movement of the bead-monomer complexes. Although a single-headed myosin-VI was able to move processively with a large cargo, the travel distance was rather short. Multiple molecules may be involved in the cargo transport for a long travel distance in cells.
authors.affiliation: Department of Biophysical Engineering, Osaka University, Suita, Osaka, Japan.
authors.forename: Mitsuhiro
authors.initials: M
authors.lastname: Iwaki
authors.forename: Hiroto
authors.initials: H
authors.lastname: Tanaka
authors.forename: Atsuko Hikikoshi
authors.initials: AH
authors.lastname: Iwane
authors.forename: Eisaku
authors.initials: E
authors.lastname: Katayama
authors.forename: Mitsuo
authors.initials: M
authors.lastname: Ikebe
authors.forename: Toshio
authors.initials: T
authors.lastname: Yanagida
mesh_terms: D000199:Actins / Q000737:chemistry* / Q000648:ultrastructure*; D001665:Binding Sites; D020409:Molecular Motor Proteins / Q000737:chemistry*; D009038:Motion; D018995:Myosin Heavy Chains / Q000737:chemistry* / Q000648:ultrastructure*; D011485:Protein Binding; D011487:Protein Conformation; D021381:Protein Transport; D013314:Stress, Mechanical
chemical_list: D000199:Actins; D020409:Molecular Motor Proteins; C089785:myosin VI; D018995:Myosin Heavy Chains