Abstract
Cell movement and cytokinesis are facilitated by contractile forces generated by the molecular motor, non-muscle myosin II (NMII). NMII molecules form a filament (NMII-F) through interactions of their C-terminal rod domains, positioning groups of N-terminal motor domains on opposite sides. The NMII motors then bind and pull actin filaments toward the NMII-F, thus driving contraction. Inside of crawling cells, NMIIA-Fs form large macromolecular ensembles (i.e., NMIIA-F stacks) but how this occurs is unknown. Here we show NMIIA-F stacks are formed through two non-mutually exclusive mechanisms: expansion and concatenation. During expansion, NMIIA molecules within the NMIIA-F spread out concurrent with addition of new NMIIA molecules. Concatenation occurs when multiple NMIIA-F/NMIIA-F stacks move together and align. We found NMIIA-F stack formation was regulated by both motor-activity and the availability of surrounding actin filaments. Furthermore, our data showed expansion and concatenation also formed the contractile ring in dividing cells. Thus, interphase and mitotic cells share similar mechanisms for creating large contractile units, and these are likely to underlie how other myosin II-based contractile systems are assembled.