How Do Myosin VI and Myosin Va Navigate Intersections And Cooperate On Actin Tracks While Transporting Cargo In Vitro?

Abstract

Myosin Va (myoVa) and myosin VI (myoVI) are processive molecular motors that transport cargo on actin tracks in opposite directions. We have shown that myoVa can effectively maneuver through an in vitro cytoskeletal model system composed of actin filament intersections and Arp2/3 branches (Ali et al. 2007). Here we challenge Quantum dot (Qdot)-labeled expressed myoVI with actin filament intersections and observed that myoVI maneuvers through intersections with the following statistics: 38% turned left or right with equal probability; 28% crossed over the intersecting actin filament; 34% terminated their run. The myoVI cross over probability is twice that of myoVa suggesting that the range of the myoVI leading head's diffusional search may be longer than myoVa. Similar to myoVa, myoVI has significant flexibility allowing it to turn at intersection angles up to 155°. When multiple myoVI were attached a Qdot, the turning probability increased to 53% whereas the cross over probability decreased to 15%. MyoVa and myoVI may be colocalized to the same cargo in vivo and to determine how these oppositely directed motors might interact during cargo transport, we attached both motors in a 1:1 ratio to a Qdot. We observed two types of movement associated with these myoVa/myoVI-labeled Qdots. A given Qdot would move in both the plus- or minus-end direction for periods of time at velocities appropriate for the specific motor, suggesting that myoVa and myoVI take turns transporting the Qdot. Other Qdots moved continuously but at velocities suggesting that both motors are simultaneous interacting with actin and undergoing an effective “tug of war.” These studies may help characterize how actin-based motors deliver their cargo through the complex actin network.