Asymmetric Motility Powered by Myo1c on Lipid Membranes

Abstract

Class-I myosins are single-headed motors that link cell membranes to the underlying actin cytoskeleton. Actin binding occurs via the motor domain, while membrane binding has been proposed to occur via the tail domain that, in some isoforms, has been shown to interact with phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) through a putative pleckstrin homology domain. To study the interplay between myosin-I, actin, and membranes, we reconstituted actin gliding motility on a membrane using full-length vertebrate myo1c bound to fluid (D ∼ 1 μm2/s) supported lipid bilayers (SLBs) composed of 2% PtdIns(4,5)P2, 97.9% DOPC, and 0.1% TRITC-PE. In this system, myo1c dynamically attaches and detaches to/from the lipid bilayer and actin filaments. The actin-gliding velocity on SLBs at 22 °C (∼ 20 nm/s) is comparable to that of myo1c rigidly anchored to a non-fluid surface via a tail-binding monoclonal antibody. Strikingly, the gliding of actin filaments on SLBs occurs along curved paths in a counterclockwise fashion (i.e., the actin filaments turn left) when viewed from the side of the membrane. This striking asymmetric motility was not observed when the myo1c was rigidly anchored to the surface via the antibody. The tail domain was not required for filament turning, as asymmetric motility was observed with a motor domain construct (no tail) attached to fluid biotinylated SLBs via a biotin-streptavidin linkage. A slight leftward bias was also observed when full-length myo1c dynamically interacts to Ins(1,4,5)P3 that is attached to the coverslip by a flexible 3-carbon linker. We conclude that class-I myosins can produce asymmetric motility on surfaces to which they can dynamically reorient. This asymmetric motility appears to be a fundamental property of the motor domain.