Motile properties of nonmuscle myosin 2C

Nonmuscle myosin 2 (NM2) paralogs are members of actomyosin cytoskeletal complex where they hydrolyze ATP to convert chemical energy into mechanical functions in cells. NM2 are expressed in all mammalian cells where they contribute to many subcellular functions including cell migration, adhesion and cytokinesis. NM2 has three paralogs, namely NM2A, B and C. Both NM2A and NM2B are well investigated regards to their roles in disease and development with extensive characterization of their biochemical, kinetic and structural properties. NM2C, on the other hand, is the most recently identified isoform and less is known about its roles, biochemical and kinetic properties. NM2C is involved in disease and development such as, sensorineural hearing impairment and peripheral neuropathy. NM2C has roles in cell dynamics, e.g. formation of sarcomere like architecture in the tricellular apical junction and control of microvilli growth. Such roles of NM2C in cells are critically dependent on NM2C generated motility. However, the motile properties of NM2C is not studied in detail like NM2A or NM2B. Thus, study of the motile properties of NM2C will bring information that will be required to understand its subcellular function in greater detail. Here, we study the motile properties of NM2C e.g. speed at which it translocates actin, processive movement of NM2C filaments on actin and on actin-fascin bundles. We find from the in vitro motility assays that NM2C translocates actin at 46.1±0.4 (mean ± SEM) nm/s which is slowest among three NM2 paralogues. From single-filament assays, we also show that NM2C filaments move processively on both actin (27.8±0.45 nm/s) and actin-fascin bundles (22.1±0.86 nm/s). We find that NM2C filaments makes large stacks in vitro which is consistent with recent observation in cells. We want to characterize the processive movement of NM2C filaments and stacks using fluorescent probes.