Tension formation of a reconstructed nonmuscle sarcomere

The interactions between actin and nonmuscle myosin 2 enable tension dependent processes in cells such as migration and adhesion. Although cortical actomyosin networks have been described as mostly disordered, several recent super resolution light microscopy studies have shown evidence for sarcomeric arrangement of actin and myosin in various cell types. Despite their resemblance to muscle sarcomeres where bipolar myosin filaments contract antiparallel actin fibers to bring about shortening, these structures are not as easily amenable to mechanical studies due to their subcellular scale and localization. To address this gap, we reconstituted a synthetic sarcomere using laser guided micropatterning of the actin elongator formin to polymerize antiparallel actin bundles. By adding nonmuscle 2 paralogs, the tension formation and shortening of these arrangements was observed using TIRF microscopy and quantified by tracking fluorescently labeled myosin filaments. Additionally, a FRET based force-sensor was added to the myosin tail and formin, elucidating the tension formation in actin and myosin filaments. Both actin and myosin displayed features resembling isometric tension like force generation within minutes after the start of the experiment, while shortening and processive motility of myosin could be observed more than 1 hour later. This allows us to propose a mechanism of tension formation inside cells, where isometric tension of 2 pNs is mediated by sliding individual actin filaments within a network of bundles, pushing against their bundling and independently of their length. This force generation mechanism at the myosin filament level would enable cells to mediate tension and contractility over the entire cortex, while deformations occur much slower, such as during the formation of a cytokinetic ring.