Stretching Actin Filaments within Cells Enhances their Affinity for the Myosin II Motor Domain

Abstract

To test the hypothesis that the myosin II motor domain (S1) preferentially binds to specific subsets of actin filaments in vivo, we expressed GFP-fused S1 with mutations that enhanced its affinity for actin in Dictyostelium cells. Consistent with the hypothesis, the GFP-S1 mutants were localized along specific portions of the cell cortex. Comparison with rhodamine-phalloidin staining in fixed cells demonstrated that the GFP-S1 probes preferentially bound to actin filaments in the rear cortex and cleavage furrows, where actin filaments are stretched by interaction with endogenous myosin II filaments. The GFP-S1 probes were similarly enriched in the cortex stretched passively by traction forces in the absence of myosin II or by external forces using a microcapillary. The preferential binding of GFP-S1 mutants to stretched actin filaments did not depend on cortexillin I or PTEN, two proteins previously implicated in the recruitment of myosin II filaments to stretched cortex. These results suggested that it is the stretching of the actin filaments itself that increases their affinity for the myosin II motor domain. In contrast, the GFP-fused myosin I motor domain did not localize to stretched actin filaments, which suggests different preferences of the motor domains for different structures of actin filaments play a role in distinct intracellular localizations of myosin I and II. We propose a scheme in which the stretching of actin filaments, the preferential binding of myosin II filaments to stretched actin filaments, and myosin II-dependent contraction form a positive feedback loop that contributes to the stabilization of cell polarity and to the responsiveness of the cells to external mechanical stimuli.

An actin–myosin-II interaction is involved in maintaining the contractile ring in fission yeast

“…One possible mechanism for this is the preferential binding of myosin-II motor domains to mechanically stretched F-actin (Uyeda et al, 2011). Myo3 was hardly detectable on precursor medial reticular F-actin, whereas it accumulated along the sharp actin ring, which implies that the configuration or tension of the equatorial F-actin affects its affinity for Myo3.…”

Talin couples the actomyosin cortex to the plasma membrane during rear retraction and cytokinesis

“…Thus, talin A appears to bind to cortical actin filaments that are interacting with myosin II. Uyeda et al (29) proposed that the head domain of myosin II preferentially binds to actin filaments stretched by interaction with myosin II and that this binding property of the head domain is responsible for the accumulation of myosin II itself in the posterior cortex of migrating cells and cleavage furrow of dividing cells. Washington and Knecht (30) reported that the actin-binding domains of filamin and α-actinin determine the distribution of the proteins, which show distinct, almost complementary localization patterns, suggesting that different actin-binding domains have different affinities for actin filaments in functionally distinct regions of the cytoskeleton.…”

Mechanochemical Signaling Directs Cell-Shape Change

“…In mammalian nonmuscle myosin IIB, for example, when a myosin II head imposes a piconewtonrange resistive load on another, the second head releases ADP at a 10-fold slower rate than an unloaded head (0.023 5 0.003 s À1 vs. 0.27 5 0.06 s À1 ) (42). Second, in addition to inhibiting ADP release, force can trap the myosin II motor in the cooperative isometric state, which promotes the binding of additional myosin motors to the actin filament nearby due to a propagated conformational change in the actin filament (43)(44)(45). This cooperative binding state was specifically implicated in mechanosensitive accumulation by experiments in which the myosin II lever arm was lengthened or shortened (40).…”