Interacting Behavior of Two Myosin Va Motors Coupled via a DNA Scaffold

Abstract

The number of processive motors attached to a cellular cargo influences its transport behavior. Altering either motor number or the ratio of different classes of motors can therefore be a mechanism to regulate intracellular transport. Jamison et al. recently showed that two kinesin-1 motors coupled by a DNA scaffold have transport properties that are often dominated by one of the motors. Here we perform a similar experiment with myosin Va (myoVa), which has a larger step size (∼36nm) and walks on a smaller track than kinesin. A heterodimeric myoVa was labeled on only one head with either a red or green quantum dot (Qdot). Two myoVa molecules were then linked to an ∼50 nm long double-stranded DNA scaffold. Only complexes with one red and one green Qdot were analyzed. Our results show that the complex has increased run length (∼1μm) compared to a single myoVa (∼0.6μm). Average run lengths are, however, smaller than those predicted for two myosins assuming motor stepping, binding, and detachment is unaffected by intermotor interactions. Furthermore, the motor complex moved with reduced velocity (0.19 μm/s versus 0.27 μm/s for the single motor case). A histogram of the distances between the labeled heads of the two motors contains multiple peaks at ∼85, 130 and 165 nm, indicating the system is flexible. The distance between motors changes in time and the stepping pattern of the two motors are variable, suggesting asynchronous motor stepping. After the first motor binds to actin, the second motor binds at ∼10 s-1. Our findings suggest that the walking behavior of two myoVa molecules is altered when they are coupled mechanically, but perhaps in a different way than multiple kinesins. Our technique constitutes a unique tool to understand collective motor behavior.