Shrec Measurement of Myosin-VI Stepping Motion

Abstract

Myosin-VI is a motor protein that plays an important role in a large variety of cellular events such as vesicle transport and anchoring actin bundles to the plasma membrane. Myosin-VI is thought to move processively as a dimer along an actin filament in a hand-over-hand fashion with large steps similar to myosin-V. However, unlike myosin-V, its step size is largely variable. Recently, we showed using FIONA method that myosin-VI does not have widely distributed step but rather has two step types, a regular large step (72nm) and short step (44nm).(Arimoto et al. Biophysical J. vol 96 p.139a) The large steps were consistent with the hand-over-hand model. The short steps, however, were not explained by canonical stepping model. We also showed that the fraction of short steps largely increases in the presence of ADP, suggesting the short and large steps are regurated in the ADP-dependent manner. In this study, in order to investigate the coordination of two heads during short and large steps, we performed an advanced multi-color FIONA technique called single-molecule high-resolution colocalization (SHREC) which involves labeling the two heads with differently-colored Q-dots. Now we are analyzing what is the condition that myosin-VI switches between two types of stepping manner. Furthermore, to clarify how myosin-VI switches between short and long steps, we are measuring myosin-VI movement under several ADP concentrations.