Mechanosensing through cooperative interactions between myosin II and the actin crosslinker cortexillin I.

BACKGROUND: Mechanosensing governs many processes from molecular to organismal levels, including during cytokinesis where it ensures successful and symmetrical cell division. Although many proteins are now known to be force sensitive, myosin motors with their ATPase activity and force-sensitive mechanical steps are well poised to facilitate cellular mechanosensing. For a myosin motor to experience tension, the actin filament must also be anchored. RESULTS: Here, we find a cooperative relationship between myosin II and the actin crosslinker cortexillin I where both proteins are essential for cellular mechanosensory responses. Although many functions of cortexillin I and myosin II are dispensable for cytokinesis, all are required for full mechanosensing. Our analysis demonstrates that this mechanosensor has three critical elements: the myosin motor where the lever arm acts as a force amplifier, a force-sensitive bipolar thick-filament assembly, and a long-lived actin crosslinker, which anchors the actin filament so that the motor may experience tension. We also demonstrate that a Rac small GTPase inhibits this mechanosensory module during interphase, allowing the module to be primarily active during cytokinesis. CONCLUSIONS: Overall, myosin II and cortexillin I define a cellular-scale mechanosensor that controls cell shape during cytokinesis. This system is exquisitely tuned through the enzymatic properties of the myosin motor, its lever arm length, and bipolar thick-filament assembly dynamics. The system also requires cortexillin I to stably anchor the actin filament so that the myosin motor can experience tension. Through this cross-talk, myosin II and cortexillin I define a cellular-scale mechanosensor that monitors and corrects shape defects, ensuring symmetrical cell division.

Modeling cellular deformations using the level set formalism.

BACKGROUND: Many cellular processes involve substantial shape changes. Traditional simulations of these cell shape changes require that grids and boundaries be moved as the cell's shape evolves. Here we demonstrate that accurate cell shape changes can be recreated using level set methods (LSM), in which the cellular shape is defined implicitly, thereby eschewing the need for updating boundaries. RESULTS: We obtain a viscoelastic model of Dictyostelium cells using micropipette aspiration and show how this viscoelastic model can be incorporated into LSM simulations to recreate the observed protrusion of cells into the micropipette faithfully. We also demonstrate the use of our techniques by simulating the cell shape changes elicited by the chemotactic response to an external chemoattractant gradient. CONCLUSION: Our results provide a simple but effective means of incorporating cellular deformations into mathematical simulations of cell signaling. Such methods will be useful for simulating important cellular events such as chemotaxis and cytokinesis.

Interactions between myosin and actin crosslinkers control cytokinesis contractility dynamics and mechanics.

INTRODUCTION: Contractile networks are fundamental to many cellular functions, particularly cytokinesis and cell motility. Contractile networks depend on myosin-II mechanochemistry to generate sliding force on the actin polymers. However, to be contractile, the networks must also be crosslinked by crosslinking proteins, and to change the shape of the cell, the network must be linked to the plasma membrane. Discerning how this integrated network operates is essential for understanding cytokinesis contractility and shape control. Here, we analyzed the cytoskeletal network that drives furrow ingression in Dictyostelium. RESULTS: We establish that the actin polymers are assembled into a meshwork and that myosin-II does not assemble into a discrete ring in the Dictyostelium cleavage furrow of adherent cells. We show that myosin-II generates regional mechanics by increasing cleavage furrow stiffness and slows furrow ingression during late cytokinesis as compared to myoII nulls. Actin crosslinkers dynacortin and fimbrin similarly slow furrow ingression and contribute to cell mechanics in a myosin-II-dependent manner. By using FRAP, we show that the actin crosslinkers have slower kinetics in the cleavage furrow cortex than in the pole, that their kinetics differ between wild-type and myoII null cells, and that the protein dynamics of each crosslinker correlate with its impact on cortical mechanics. CONCLUSIONS: These observations suggest that myosin-II along with actin crosslinkers establish local cortical tension and elasticity, allowing for contractility independent of a circumferential cytoskeletal array. Furthermore, myosin-II and actin crosslinkers may influence each other as they modulate the dynamics and mechanics of cell-shape change.

A mechanosensory system controls cell shape changes during mitosis.

Essential life processes are heavily controlled by a variety of positive and negative feedback systems. Cytokinesis failure, ultimately leading to aneuploidy, is appreciated as an early step in tumor formation in mammals and is deleterious for all cells. Further, the growing list of cancer predisposition mutations includes a number of genes whose proteins control mitosis and/or cytokinesis. Cytokinesis shape control is also an important part of pattern formation and cell-type specialization during multi-cellular development. Inherently mechanical, we hypothesized that mechanosensing and mechanical feedback are fundamental for cytokinesis shape regulation. Using mechanical perturbation, we identified a mechanosensory control system that monitors shape progression during cytokinesis. In this review, we summarize these findings and their implications for cytokinesis regulation and for understanding the cytoskeletal system architecture that governs shape control.

Mitosis-specific mechanosensing and contractile-protein redistribution control cell shape.

Because cell-division failure is deleterious, promoting tumorigenesis in mammals, cells utilize numerous mechanisms to control their cell-cycle progression. Though cell division is considered a well-ordered sequence of biochemical events, cytokinesis, an inherently mechanical process, must also be mechanically controlled to ensure that two equivalent daughter cells are produced with high fidelity. Given that cells respond to their mechanical environment, we hypothesized that cells utilize mechanosensing and mechanical feedback to sense and correct shape asymmetries during cytokinesis. Because the mitotic spindle and myosin II are vital to cell division, we explored their roles in responding to shape perturbations during cell division. We demonstrate that the contractile proteins myosin II and cortexillin I redistribute in response to intrinsic and externally induced shape asymmetries. In early cytokinesis, mechanical load overrides spindle cues and slows cytokinesis progression while contractile proteins accumulate and correct shape asymmetries. In late cytokinesis, mechanical perturbation also directs contractile proteins but without apparently disrupting cytokinesis. Significantly, this response only occurs during anaphase through cytokinesis, does not require microtubules, and is independent of spindle orientation, but is dependent on myosin II. Our data provide evidence for a mechanosensory system that directs contractile proteins to regulate cell shape during mitosis.

Enlazin, a natural fusion of two classes of canonical cytoskeletal proteins, contributes to cytokinesis dynamics.

Cytokinesis requires a complex network of equatorial and global proteins to regulate cell shape changes. Here, using interaction genetics, we report the first characterization of a novel protein, enlazin. Enlazin is a natural fusion of two canonical classes of actin-associated proteins, the ezrin-radixin-moesin family and fimbrin, and it is localized to actin-rich structures. A fragment of enlazin, enl-tr, was isolated as a genetic suppressor of the cytokinesis defect of cortexillin-I mutants. Expression of enl-tr disrupts expression of endogenous enlazin, indicating that enl-tr functions as a dominant-negative lesion. Enlazin is distributed globally during cytokinesis and is required for cortical tension and cell adhesion. Consistent with a role in cell mechanics, inhibition of enlazin in a cortexillin-I background restores cytokinesis furrowing dynamics and suppresses the growth-in-suspension defect. However, as expected for a role in cell adhesion, inhibiting enlazin in a myosin-II background induces a synthetic cytokinesis phenotype, frequently arresting furrow ingression at the dumbbell shape and/or causing recession of the furrow. Thus, enlazin has roles in cell mechanics and adhesion, and these roles seem to be differentially significant for cytokinesis, depending on the genetic background.

The stress and strain of cytokinesis.

The ultimate goal of all signaling pathways in cytokinesis is to control the mechanical separation of the mother cell into two daughter cells. Because of the intrinsic mechanical nature of cytokinesis, it is essential to understand fully how cell shapes and the material properties of the cell are generated, how these shapes and material properties create force, and how motor proteins such as myosin-II modify the system to achieve successful cytokinesis. In this review (which is part of the Cytokinesis series), we discuss the relevant physical properties of cells, how these properties are measured and the basic models that are used to understand cell mechanics. Finally, we present our current understanding of how cytokinesis mechanics work.