Myosin-18B Promotes the Assembly of Myosin II Stacks for Maturation of Contractile Actomyosin Bundles

Cell adhesion, morphogenesis, mechanosensing, and muscle contraction rely on contractile actomyosin bundles, where the force is produced through sliding of bipolar myosin II filaments along actin filaments. The assembly of contractile actomyosin bundles involves registered alignment of myosin II filaments and their subsequent fusion into large stacks. However, mechanisms underlying the assembly of myosin II stacks and their physiological functions have remained elusive. Here, we identified myosin-18B, an unconventional myosin, as a stable component of contractile stress fibers. Myosin-18B co-localized with myosin II motor domains in stress fibers and was enriched at the ends of myosin II stacks. Importantly, myosin-18B deletion resulted in drastic defects in the concatenation and persistent association of myosin II filaments with each other and thus led to severely impaired assembly of myosin II stacks. Consequently, lack of myosin-18B resulted in defective maturation of actomyosin bundles from their precursors in osteosarcoma cells. Moreover, myosin-18B knockout cells displayed abnormal morphogenesis, migration, and ability to exert forces to the environment. These results reveal a critical role for myosin-18B in myosin II stack assembly and provide evidence that myosin II stacks are important for a variety of vital processes in cells. [Display omitted] •Myosin-18B is a stable component of stress fibers in osteosarcoma cells•Stress-fiber maturation from their precursors requires myosin-18B•Myosin-18B promotes the assembly of myosin II stacks•Myosin II stacks are important for cell morphogenesis and migration Jiu et al. demonstrate that a highly divergent member of the myosin family, myosin-18B, is critical for assembly of myosin II stacks in human osteosarcoma cells. They also provide evidence that these higher-order myosin II structures are important for generation of functional actomyosin bundles in cells.