Molecular Characterization and Subcellular Localization of Arabidopsis class VIII myosin, ATM1 [Plant Biology]

March 17th, 2014 by Haraguchi, T., Tominaga, M., Matsumoto, R., Sato, K., Nakano, A., Yamamoto, K., Ito, K.

Land plants possess myosin classes VIII and XI. Although some information is available on the molecular properties of class XI myosins, class VIII myosins are not characterized. Here we report the first analysis of the enzymatic properties of class VIII myosin. The motor domain of Arabidopsis class VIII myosin, ATM1 (ATM1-MD) and the motor domain plus one IQ motif (ATM1-1IQ) were expressed in a baculovirus system and characterized. ATM1-MD and ATM1-1IQ had low actin-activated Mg2+-ATPase activity (Vmax = 4 s−1), although their affinities for actin were high (Kactin = 4 μM). The actin-sliding velocities of ATM1-MD and ATM1-1IQ were 0.02 and 0.089 μm/s, respectively, from which the value for full-length ATM1 is calculated to be approximately 0.2 μm/s. The results of actin co-sedimentation assay showed that duty ratio of ATM1 was approximately 90%. ADP dissociation from the actin-ATM1 complex (acto-ATM1) was extremely slow, which accounts for the low actin-sliding velocity, low actin-activated ATPase activity, and high duty ratio. The rate of ADP dissociation from acto-ATM1 was markedly biphasic with fast- and slow-phase rates (5.1 s−1 and 0.41 s−1, respectively). Physiological concentrations of free Mg 2+ modulated actin-sliding velocity and actin-activated ATPase activity by changing the rate of ADP dissociation from acto-ATM1. GFP-fused full-length ATM1 expressed in Arabidopsis was localized to actin filaments at the cell cortex, plasmodesmata, plastids, and. newly formed cell walls. Our results suggest that ATM1 functions as a tension sensor/generator at the cell cortex and other structures in Arabidopsis.