Spatial integration of mechanical forces by α-actinin establishes actin network symmetry

Senger, Fabrice; Pitaval, Amandine; Ennomani, Hajer; Kurzawa, Laëtitia; Blanchoin, Laurent; Théry, Manuel

doi:10.1242/jcs.236604

Cited by 35 publications

(24 citation statements)

References 62 publications

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“…S4H) as well as the spatial correlation of traction force vectors (SI Appendix, Fig. S4I) was highest in ACTN4-EGFP overexpressing cells and decreased as ACTN cross-linking was reduced, which implied that the spatial symmetry of traction forces was controlled by ACTN concentration, in line with another study (28).…”

Section: Rigidity-dependent Cell Polarization Is Tuned By Passive Actnsupporting

confidence: 85%

“…In vitro rheological studies on actin gels show a power-law type relationship between elastic modulus and cross-linker concentration (23) which depends on the specific binding affinity between the cross-linker and actin (24), the strain rate (25), and the prestress (26). Global actomyosin contraction dynamics are directly regulated by the amount of ACTN cross-linking in an actin gel (27) and lack of ACTN has been also shown to disrupt actin network symmetry in vitro and in cells (28). Here, we describe a functional role of myosin II and ACTN cross-linking in preserving cell symmetry, regulating cell-scale ordering of the cytoskeleton, and tuning the cytoskeleton's adaptive response to substrate rigidity that reflects changes in adhesions incurred via local rigidity sensing.…”

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confidence: 99%

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Cell response to substrate rigidity is regulated by active and passive cytoskeletal stress

Doss

Meng

Gupta

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

143

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Morphogenesis, tumor formation, and wound healing are regulated by tissue rigidity. Focal adhesion behavior is locally regulated by stiffness; however, how cells globally adapt, detect, and respond to rigidity remains unknown. Here, we studied the interplay between the rheological properties of the cytoskeleton and matrix rigidity. We seeded fibroblasts onto flexible microfabricated pillar arrays with varying stiffness and simultaneously measured the cytoskeleton organization, traction forces, and cell-rigidity responses at both the adhesion and cell scale. Cells adopted a rigidity-dependent phenotype whereby the actin cytoskeleton polarized on stiff substrates but not on soft. We further showed a crucial role of active and passive cross-linkers in rigidity-sensing responses. By reducing myosin II activity or knocking down α-actinin, we found that both promoted cell polarization on soft substrates, whereas α-actinin overexpression prevented polarization on stiff substrates. Atomic force microscopy indentation experiments showed that this polarization response correlated with cell stiffness, whereby cell stiffness decreased when active or passive cross-linking was reduced and softer cells polarized on softer matrices. Theoretical modeling of the actin network as an active gel suggests that adaptation to matrix rigidity is controlled by internal mechanical properties of the cytoskeleton and puts forward a universal scaling between nematic order of the actin cytoskeleton and the substrate-to-cell elastic modulus ratio. Altogether, our study demonstrates the implication of cell-scale mechanosensing through the internal stress within the actomyosin cytoskeleton and its coupling with local rigidity sensing at focal adhesions in the regulation of cell shape changes and polarity.

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Section: Rigidity-dependent Cell Polarization Is Tuned By Passive Actnsupporting

confidence: 85%

mentioning

confidence: 99%

Cell response to substrate rigidity is regulated by active and passive cytoskeletal stress

Doss

Meng

Gupta

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

143

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“…2a) 32 . Each pattern elicited a distinguishable actin 3D architecture 26,33,34 , with spatially distinct peripheral and internal stress fibers, transverse arcs and isotropic branched meshworks. To elucidate the actin organization underlying these architectures, we first performed cellular tomography directly on the thin peripheries of micropattern-shaped cells (Fig.…”

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confidence: 99%

Tailoring cryo-electron microscopy grids by photo-micropatterning for in-cell structural studies

et al. 2019

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Spatially-controlled cell adhesion on electron microscopy (EM) supports remains a bottleneck in specimen preparation for cellular cryo-electron tomography. Here, we describe contactless and mask-free photo-micropatterning of EM grids for site-specific deposition of extracellular matrix (ECM)-related proteins. We attained refined cell positioning for micromachining by cryo-focused ion beam milling. Complex micropatterns generated predictable intracellular organization, allowing direct correlation between cell architecture and in-cell 3D-structural characterization of the underlying molecular machinery. In parallel to the ongoing resolution revolution in cryo-electron microscopy for macromolecular structure determination 1 , cryo-electron tomography (ET) has matured to reveal the molecular sociology in situ sensu stricto 2-4. Yet, molecular-resolution cryo-ET of adherent mammalian cells can only be directly performed on their thin peripheries (< 300 nm). To reveal structures at the cell interior, thinning by cryo-focused ion beam (FIB) has proved an optimal, artifact-free preparation method 2,5-12. Thus, cryo-FIB micromachining coupled to cryo-ET, followed by subtomogram averaging 13,14 , allows the capture of Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:

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“…This underscores the importance of network connectivity by α-actinin in regulating actin network contraction and possibly aggregation. In reconstituted actomyosin networks on circular micropatterns, contraction induced the formation of star-like patterns and the presence of α-actinin resulted in centering of the star-like patterns 44 . α-actinin also mediated droplet-size-dependent cluster localization of encapsulated actomyosin networks 45 .…”

Section: Encapsulated α-Actinin and Fascin Together Form Distinct Actmentioning

confidence: 99%

“…α-actinin also mediated droplet-size-dependent cluster localization of encapsulated actomyosin networks 45 . In cells geometrically confined on micropatterned substrates, α-actinin was shown to modulate symmetric actomyosin-driven centering 44 . Hence, α-actinin-regulated network connectivity and the possibility of a GUV size-dependent clustering directed us to explore the role of confinement and α-actinin-mediated self-organization in aggregation and architecture of actin assemblies.…”

Section: Encapsulated α-Actinin and Fascin Together Form Distinct Actmentioning

confidence: 99%

Actin crosslinker competition and sorting drive emergent GUV size-dependent actin network architecture

Bashirzadeh

Redford

Lorpaiboon

et al. 2020

Preprint

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Robust spatiotemporal organization of cytoskeletal networks is crucial, enabling cellular processes such as cell migration and division. α-Actinin and fascin are two actin crosslinking proteins localized to distinct regions of eukaryotes to form actin bundles with optimized spacing for cell contractile machinery and sensory projections, respectively. In vitro reconstitution assays and coarse-grained simulations have shown that these actin bundling proteins segregate into distinct domains with a bundler size-dependent competition-based mechanism, driven by the minimization of F-actin bending energy. However, it is not known how physical confinement imposed by the cell membrane contributes to sorting of actin bundling proteins and the concomitant reorganization of actin networks in intracellular environment. Here, by encapsulating actin, α-actinin, and fascin in giant unilamellar vesicles (GUVs), we show that the size of such a spherical boundary determines equilibrated structure of actin networks among three typical structures: single rings, astral structures, and star-like structures. We show that α-actinin bundling activity and its tendency for clustering actin is central to the formation of these structures. By analyzing physical features of crosslinked actin networks, we show that spontaneous sorting and domain formation of α-actinin and fascin are intimately linked to the resulting structures. We propose that the observed boundary-imposed effect on sorting and structure formation is a general mechanism by which cells can select between different structural dynamical steady states.

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Spatial integration of mechanical forces by α-actinin establishes actin network symmetry

Cited by 35 publications

References 62 publications

Cell response to substrate rigidity is regulated by active and passive cytoskeletal stress

Cell response to substrate rigidity is regulated by active and passive cytoskeletal stress

Tailoring cryo-electron microscopy grids by photo-micropatterning for in-cell structural studies

Actin crosslinker competition and sorting drive emergent GUV size-dependent actin network architecture

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