Small teams of myosin Vc coordinate their stepping for efficient cargo transport on actin bundles

Myosin Vc (myoVc) is unique among vertebrate class V myosin isoforms in that it requires teams of motors to transport cargo. Single molecules of myoVc cannot take multiple steps on single actin filaments, in stark contrast to the well-studied myosin Va (myoVa) isoform. Consistent with in vivo studies (1), only teams of myoVc motors can move continuously on actin bundles at physiologic ionic strength (2), raising the question of how motor-motor interactions cause this preference. Here, using DNA nanostructures as synthetic cargos for linking defined numbers of myoVa or myoVc molecules, we compared the stepping behavior of myoVa versus myoVc teams, and myoVc stepping patterns on single actin filaments versus actin bundles. Run lengths of both myoVa and myoVc teams increased with motor number, but the run lengths of myoVc teams were longer on actin bundles than on filaments. By resolving the stepping behavior of individual myoVc motors with a Qdot bound to the motor domain, we found that coupling of two myoVc molecules significantly decreases futile back/side steps, which were frequently observed for single myoVc motors. Data showing how changes in the inter-motor distance of two coupled myoVc motors affected stepping dynamics suggested that mechanical tension coordinates the stepping behavior of two molecules for efficient directional motion. Our study thus provides a molecular basis to explain how teams of myoVc motors are suited to transport cargos such as zymogen granules on actin bundles.