scholarly journals Using optogenetics to link myosin patterns to contractile cell behaviors during convergent extension

2021 ◽  
Author(s):  
R. M. Herrera-Perez ◽  
C. Cupo ◽  
C. Allan ◽  
A. Lin ◽  
K. E. Kasza
Keyword(s):  
Myosin Ii ◽  
Apical Cell ◽  
Epithelial Tissue ◽  
Cell Area ◽  
Convergent Extension ◽  
Cell Behaviors ◽  
Actomyosin Contractility ◽  
Axis Elongation ◽  
Cell Intercalation ◽  
Shape Changes

ABSTRACTDistinct spatiotemporal patterns of actomyosin contractility are often associated with particular epithelial tissue shape changes during development. For example, a planar polarized pattern of myosin II localization regulated by Rho1 signaling duringDrosophilabody axis elongation is thought to drive the cell behaviors that contribute to convergent extension. However, it is not well understood how specific aspects of a myosin localization pattern influence the multiple cell behaviors—including cell intercalation, cell shape changes, and apical cell area fluctuations—that simultaneously occur within a tissue during morphogenesis. Here, we use optogenetic activation (optoGEF) and deactivation (optoGAP) of Rho1 signaling to perturb the myosin pattern in the germband epithelium duringDrosophilaaxis elongation and analyze the effects on contractile cell behaviors within the tissue. We find that uniform photoactivation of optoGEF or optoGAP is sufficient to rapidly override the endogenous myosin pattern, abolishing myosin planar polarity and reducing cell intercalation and convergent extension. However, these two perturbations have distinct effects on junctional and medial myosin localization, apical cell area fluctuations, and cell packings within the germband. Activation of Rho1 signaling in optoGEF embryos increases myosin accumulation in the medial-apical domain of germband cells, leading to increased amplitudes of apical cell area fluctuations. This enhanced contractility is translated into heterogeneous reductions in apical cell areas across the tissue, disrupting cellular packings within the germband. Conversely, inactivation of Rho1 signaling in optoGAP embryos decreases both medial and junctional myosin accumulation, leading to a dramatic reduction in cell area fluctuations. These results demonstrate that the level of Rho1 activity and the balance between junctional and medial myosin regulate apical cell area fluctuations and cellular packings in the germband, which have been proposed to influence the biophysics of cell rearrangements and tissue fluidity.STATEMENT OF SIGNIFICANCETissues are shaped by forces produced by dynamic patterns of actomyosin contractility. However, the mechanisms underlying these myosin patterns and their translation into cell behavior and tissue-level movements are not understood. Here, we show that optogenetic tools designed to control upstream regulators of myosin II can be used to rapidly manipulate myosin patterns and analyze the effects on cell behaviors during tissue morphogenesis. Combining optogenetics with live imaging in the developing fruit fly embryo, we show that acute perturbations to upstream myosin regulators are sufficient to rapidly perturb existing myosin patterns and alter cell movements and shapes during axis elongation, resulting in abnormalities in embryo shape. These results directly link myosin contractility patterns to cell behaviors that shape tissues, providing new insights into the mechanisms that generate functional tissues.

scholarly journals Optogenetic dissection of the roles of actomyosin in the mechanics underlying tissue fluidity

2021 ◽  
Author(s):  
R. Marisol Herrera-Perez ◽  
Christian Cupo ◽  
Cole Allan ◽  
Alicia B. Dagle ◽  
Karen E. Kasza
Keyword(s):  
Mechanical Properties ◽  
Rho Kinase ◽  
Tissue Level ◽  
Epithelial Tissue ◽  
Apical Surface ◽  
Convergent Extension ◽  
Actomyosin Contractility ◽  
Mechanical Tensions ◽  
Axis Elongation ◽  
Complex Relationships

Rapid epithelial tissue flows are essential to building and shaping developing embryos. However, it is not well understood how the mechanical properties of tissues and the forces driving them to flow are jointly regulated to accommodate rapid tissue remodeling. To dissect the roles of actomyosin in the mechanics of epithelial tissue flows, here we use two optogenetic tools, optoGEF and optoGAP, to manipulate Rho/Rho-kinase signaling and actomyosin contractility in the germband epithelium, which flows via convergent extension during Drosophila body axis elongation. The ability to perturb actomyosin across the tissue allows us to analyze the effects of actomyosin on cell rearrangements, tissue tensions, and tissue mechanical properties. We find that either optogenetic activation or deactivation of Rho1 signaling and actomyosin contractility at the apical surface of the germband disrupts cell rearrangements and tissue-level flows. By probing mechanical tensions in the tissue using laser ablation and predicting tissue mechanical properties from cell packings, we find that actomyosin influences both the anisotropic forces driving tissue flow and the mechanical properties of the tissue resisting flow, leading to complex relationships between actomyosin activity and tissue-level flow. Moreover, our results indicate that changes in the distribution of medial and junctional myosin in the different perturbations alter tissue tension and cell packings in distinct ways, revealing how junctional and medial myosin have differential roles in promoting and orienting cell rearrangements to tune tissue flows in developing embryos.

scholarly journals Rho GTPase and Shroom direct planar polarized actomyosin contractility during convergent extension

2014 ◽  
Vol 204 (4) ◽  
pp. 575-589 ◽  
Author(s):  
Sérgio de Matos Simões ◽  
Avantika Mainieri ◽  
Jennifer A. Zallen
Keyword(s):  
Rho Gtpase ◽  
Binding Protein ◽  
Myosin Ii ◽  
Rho Kinase ◽  
Actin Binding ◽  
Mechanical Forces ◽  
Convergent Extension ◽  
Planar Polarity ◽  
Axis Elongation ◽  
Actomyosin Contraction

Actomyosin contraction generates mechanical forces that influence cell and tissue structure. During convergent extension in Drosophila melanogaster, the spatially regulated activity of the myosin activator Rho-kinase promotes actomyosin contraction at specific planar cell boundaries to produce polarized cell rearrangement. The mechanisms that direct localized Rho-kinase activity are not well understood. We show that Rho GTPase recruits Rho-kinase to adherens junctions and is required for Rho-kinase planar polarity. Shroom, an asymmetrically localized actin- and Rho-kinase–binding protein, amplifies Rho-kinase and myosin II planar polarity and junctional localization downstream of Rho signaling. In Shroom mutants, Rho-kinase and myosin II achieve reduced levels of planar polarity, resulting in decreased junctional tension, a disruption of multicellular rosette formation, and defective convergent extension. These results indicate that Rho GTPase activity is required to establish a planar polarized actomyosin network, and the Shroom actin-binding protein enhances myosin contractility locally to generate robust mechanical forces during axis elongation.

scholarly journals Geometric control of Myosin-II orientation during axis elongation

2022 ◽  
Author(s):  
Matthew Frederick Lefebvre ◽  
Nikolas Heinrich Claussen ◽  
Noah Prentice Mitchell ◽  
Hannah J Gustafson ◽  
Sebastian J Streichan
Keyword(s):  
Gene Expression ◽  
Large Scale ◽  
Time Course ◽  
Protein Localization ◽  
Expression Patterns ◽  
Myosin Ii ◽  
Convergent Extension ◽  
Pair Rule Gene ◽  
Axis Elongation ◽  
Actomyosin Cytoskeleton

The actomyosin cytoskeleton is a crucial driver of morphogenesis. Yet how the behavior of large scale cytoskeletal patterns in deforming tissues emerges from the interplay of geometry, genetics, and mechanics remains incompletely understood. Convergent extension flow in D. melanogaster embryos provides the opportunity to establish a quantitative understanding of the dynamics of anisotropic non-muscle myosin II. Cell-scale analysis of protein localization in fixed embryos suggests that there are complex rules governing how the control of myosin anisotropy is regulated by gene expression patterns. However, technical limitations have impeded quantitative and dynamic studies of this process at the whole embryo level, leaving the role of geometry open. Here we combine in toto live imaging with quantitative analysis of molecular dynamics to characterize the distribution of myosin anisotropy and corresponding genetic patterning. We found pair rule gene expression continuously deformed, flowing with the tissue frame. In contrast, myosin anisotropy orientation remained nearly static, aligned with the stationary dorsal-ventral axis of the embryo. We propose myosin recruitment by a geometrically defined static source, potentially related to the embryo-scale epithelial tension, and account for transient deflections by the interplay of cytoskeletal turnover with junction reorientation by flow. With only one parameter, this model quantitatively accounts for the time course of myosin anisotropy orientation in wild-type, twist, and even-skipped embryos as well as embryos with perturbed egg geometry. Geometric patterning of the cytoskeleton suggests a simple physical strategy to ensure a robust flow and formation of shape.

scholarly journals Vertex sliding drives intercalation by radial coupling of adhesion and actomyosin networks during Drosophila germband extension

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Timothy E Vanderleest ◽  
Celia M Smits ◽  
Yi Xie ◽  
Cayla E Jewett ◽  
J Todd Blankenship ◽  
...  
Keyword(s):  
Developmental Process ◽  
Myosin Ii ◽  
Radial Force ◽  
Force Balance ◽  
Cell Area ◽  
Systematic Analysis ◽  
Interface Length ◽  
Cell Intercalation ◽  
E Cadherin ◽  
Cell Cell

Oriented cell intercalation is an essential developmental process that shapes tissue morphologies through the directional insertion of cells between their neighbors. Previous research has focused on properties of cell–cell interfaces, while the function of tricellular vertices has remained unaddressed. Here, we identify a highly novel mechanism in which vertices demonstrate independent sliding behaviors along cell peripheries to produce the topological deformations responsible for intercalation. Through systematic analysis, we find that the motion of vertices connected by contracting interfaces is not physically coupled, but instead possess strong radial coupling. E-cadherin and Myosin II exist in previously unstudied populations at cell vertices and undergo oscillatory cycles of accumulation and dispersion that are coordinated with changes in cell area. Additionally, peak enrichment of vertex E-cadherin/Myosin II coincides with interface length stabilization. Our results suggest a model in which asymmetric radial force balance directs the progressive, ratcheted motion of individual vertices to drive intercalation.

scholarly journals Myosin-II-mediated cell shape changes and cell intercalation contribute to primitive streak formation

2015 ◽  
Vol 17 (4) ◽  
pp. 397-408 ◽  
Author(s):  
Emil Rozbicki ◽  
Manli Chuai ◽  
Antti I. Karjalainen ◽  
Feifei Song ◽  
Helen M. Sang ◽  
...  
Keyword(s):  
Cell Shape ◽  
Myosin Ii ◽  
Primitive Streak ◽  
Cell Shape Changes ◽  
Cell Intercalation ◽  
Shape Changes

Induction of notochord cell intercalation behavior and differentiation by progressive signals in the gastrula of Xenopus laevis

Development ◽  
1995 ◽  
Vol 121 (10) ◽  
pp. 3311-3321 ◽  
Author(s):  
C. Domingo ◽  
R. Keller
Keyword(s):  
Xenopus Laevis ◽  
Behavioral Changes ◽  
Cell Behavior ◽  
Convergent Extension ◽  
Notochordal Cell ◽  
Cell Behaviors ◽  
Notochord Cell ◽  
Light Fluorescence ◽  
Cell Intercalation ◽  
Somitic Mesoderm

We show that notochord-inducing signals are present during Xenopus laevis gastrulation and that they are important for both inducing and organizing cell behavior and differentiation in the notochord. Previous work showed that convergent extension of prospective notochordal and somitic mesoderm occurs by mediolateral cell intercalation to produce a longer, narrower tissue. Mediolateral cell intercalation is driven by bipolar, mediolaterally directed protrusive activity that elongates cells and then pulls them between one another along the mediolateral axis. This cell behavior, and subsequent notochordal cell differentiation, begins anteriorly and spreads posteriorly along the notochordal-somitic boundary, and from this lateral boundary progresses medially towards the center of the notochord field. To examine whether these progressions of cell behaviors and differentiation are induced and organized during gastrulation, we grafted labeled cells from the prospective notochordal, somitic and epidermal regions of the gastrula into the notochordal region and monitored their behavior by low light, fluorescence videomicroscopy. Prospective notochordal, epidermal and somitic cells expressed mediolateral cell intercalation behavior in an anterior-to-posterior and lateral-to-medial order established by the host notochord. Behavioral changes were induced first and most dramatically among cells grafted next to the notochordal-somitic boundary, particularly those in direct contact with the boundary, suggesting that the boundary may provide signals that both induce and organize notochordal cell behaviors. By physically impeding normal convergent extension movements, notochordal cell behaviors and differentiation were restricted to the anteriormost notochordal region and to the lateral notochordal-somitic boundary. These results show that mediolateral cell intercalation behavior and notochordal differentiation can be induced in the gastrula stage, among cells not normally expressing these characteristics, and that these characteristics are induced progressively, most likely by signals emanating from the notochordal-somitic boundary. In addition, they show that morphogenetic movements during gastrulation are necessary for complete notochord formation and that the prospective notochord region is not determined by the onset of gastrulation.

scholarly journals Myosin-dependent remodeling of adherens junctions protects junctions from Snail-dependent disassembly

2016 ◽  
Vol 212 (2) ◽  
pp. 219-229 ◽  
Author(s):  
Mo Weng ◽  
Eric Wieschaus
Keyword(s):  
Cell Shape ◽  
Adherens Junctions ◽  
Myosin Ii ◽  
Cell Junctions ◽  
Snail Expression ◽  
Quantitative Image ◽  
Early Expression ◽  
Temporally Correlated ◽  
Shape Changes

Although Snail is essential for disassembly of adherens junctions during epithelial–mesenchymal transitions (EMTs), loss of adherens junctions in Drosophila melanogaster gastrula is delayed until mesoderm is internalized, despite the early expression of Snail in that primordium. By combining live imaging and quantitative image analysis, we track the behavior of E-cadherin–rich junction clusters, demonstrating that in the early stages of gastrulation most subapical clusters in mesoderm not only persist, but move apically and enhance in density and total intensity. All three phenomena depend on myosin II and are temporally correlated with the pulses of actomyosin accumulation that drive initial cell shape changes during gastrulation. When contractile myosin is absent, the normal Snail expression in mesoderm, or ectopic Snail expression in ectoderm, is sufficient to drive early disassembly of junctions. In both cases, junctional disassembly can be blocked by simultaneous induction of myosin contractility. Our findings provide in vivo evidence for mechanosensitivity of cell–cell junctions and imply that myosin-mediated tension can prevent Snail-driven EMT.

scholarly journals Unipolar distributions of junctional Myosin II identify cell stripe boundaries that drive cell intercalation throughout Drosophila axis extension

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Robert J Tetley ◽  
Guy B Blanchard ◽  
Alexander G Fletcher ◽  
Richard J Adams ◽  
Bénédicte Sanson
Keyword(s):  
Myosin Ii ◽  
Interaction Model ◽  
Cell Interaction ◽  
Cell Number ◽  
Cell Polarization ◽  
Convergence And Extension ◽  
Pair Rule Gene ◽  
Cell Intercalation ◽  
Pair Rule ◽  
Cell Cell

Convergence and extension movements elongate tissues during development. Drosophila germ-band extension (GBE) is one example, which requires active cell rearrangements driven by Myosin II planar polarisation. Here, we develop novel computational methods to analyse the spatiotemporal dynamics of Myosin II during GBE, at the scale of the tissue. We show that initial Myosin II bipolar cell polarization gives way to unipolar enrichment at parasegmental boundaries and two further boundaries within each parasegment, concomitant with a doubling of cell number as the tissue elongates. These boundaries are the primary sites of cell intercalation, behaving as mechanical barriers and providing a mechanism for how cells remain ordered during GBE. Enrichment at parasegment boundaries during GBE is independent of Wingless signaling, suggesting pair-rule gene control. Our results are consistent with recent work showing that a combinatorial code of Toll-like receptors downstream of pair-rule genes contributes to Myosin II polarization via local cell-cell interactions. We propose an updated cell-cell interaction model for Myosin II polarization that we tested in a vertex-based simulation.

scholarly journals Long-term, in toto live imaging of the developing mouse heart

2020 ◽  
Author(s):  
Yanzhu Yue ◽  
Xin Li ◽  
Youdong Zhang ◽  
Aibin He
Keyword(s):  
Imaging System ◽  
Light Sheet ◽  
Live Imaging ◽  
Mouse Heart ◽  
Cell Behaviors ◽  
Oriented Cell Division ◽  
Lower Vertebrates ◽  
Imaging Strategy ◽  
Cell Intercalation ◽  
Gated Imaging

Abstract Mapping holistic cell behaviors sculpting mammalian heart has been a goal, but so far only successes in transparent invertebrates and lower vertebrates. Using a live-imaging system comprising a customized vertical light-sheet microscope equipped with a culture module, a heartbeat-gated imaging strategy, and a digital image processing framework, we realized imaging of developing mouse hearts with uninterrupted cell lineages for up to 1.5 days. Four-dimensional landscapes of cell behaviors revealed a blueprint for ventricle chamber formation in which biased outward migration of outermost cardiomyocytes coupled with cell intercalation and horizontal division. The trabeculae, an inner muscle architecture, was developed through early fate segregation and transmural cell arrangement involving both oriented cell division and directional migration. Thus, live-imaging reconstruction affords a transformative means for deciphering mammalian organogenesis.

scholarly journals Mechanical state, material properties and continuous description of an epithelial tissue

2012 ◽  
Vol 9 (75) ◽  
pp. 2614-2623 ◽  
Author(s):  
Isabelle Bonnet ◽  
Philippe Marcq ◽  
Floris Bosveld ◽  
Luc Fetler ◽  
Yohanns Bellaïche ◽  
...  
Keyword(s):  
Continuous Model ◽  
Epithelial Tissue ◽  
Mechanical State ◽  
Continuous Description ◽  
Time Continuum ◽  
Shaped Domain ◽  
Internal Viscosity ◽  
Shape Changes ◽  
Good Agreement

During development, epithelial tissues undergo extensive morphogenesis based on coordinated changes of cell shape and position over time. Continuum mechanics describes tissue mechanical state and shape changes in terms of strain and stress. It accounts for individual cell properties using only a few spatially averaged material parameters. To determine the mechanical state and parameters in the Drosophila pupa dorsal thorax epithelium, we severed in vivo the adherens junctions around a disc-shaped domain comprising typically a hundred cells. This enabled a direct measurement of the strain along different orientations at once. The amplitude and the anisotropy of the strain increased during development. We also measured the stress-to-viscosity ratio and similarly found an increase in amplitude and anisotropy. The relaxation time was of the order of 10 s. We propose a space–time, continuous model of the relaxation. Good agreement with experimental data validates the description of the epithelial domain as a continuous, linear, visco-elastic material. We discuss the relevant time and length scales. Another material parameter, the ratio of external friction to internal viscosity, is estimated by fitting the initial velocity profile. Together, our results contribute to quantify forces and displacements, and their time evolution, during morphogenesis.

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