Comparing the Motility of Myosin X with Parallel and Anti-Parallel Dimerization Domains

Myosin X is an actin-based molecular motor involved in filopodial formation, cell migration, adhesion and mitotic spindle orientation. It is upregulated in some cancers and may play a role in metastasis. Myosin X induces filopodial formation and localizes to filopodial tips by an unclear targeting mechanism. Studies using various truncated, forced-dimers have indicated that processivity of myosin X is greater on fascin-bundled actin filaments than single filaments, whereas other studies have not found different processivity on bundles relative to single filaments. Recently, the native dimerization domain has been proposed to form anti-parallel dimers, which might influence aspects of motility such as preference for actin bundles. To test this idea, we compared processive motility of forced parallel and anti-parallel myosin X constructs on single and bundled actin filaments. On single actin filaments parallel constructs are robustly processive, while processivity of the anti-parallel construct is much weaker. Run lengths for anti-parallel myosin X decreased as [MgATP] was raised from 0.5 to 2.5 micromolar. At 5 micromolar MgATP, run lengths are shorter than our detection limit, ∼100 nm. The velocity of the anti-parallel construct on single actin filaments is slower than that of the parallel construct, due to a substantial proportion of short (e.g. +18 nm) and backward steps along with +36 nm steps. On fascin-actin bundles, however, the processivity, velocity, step-size, and backward stepping frequency are similar to those characteristics of parallel myosin X. These results suggest that in cells, the anti-parallel coiled-coil domain may help myosin X select structures containing parallel actin bundles, such as filopodia. Supported by NIH grant R01GM086352 and T32HL07954 and TEXAS STAR Plus award.