Actin cytoskeleton self-organization in single epithelial cells and fibroblasts under isotropic confinement

Jalal, Salma; Huang, Ruby Yun‐Ju; Viasnoff, Virgile; Tee, Yee Han; Bershadsky, Alexander D.

doi:10.1101/318022

Cited by 13 publications

(14 citation statements)

References 44 publications

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“…Surprisingly, when comparing between the two chondrocyte cell types, we also observed enhancement of SOX9 and COL2A1 expression on 62 nm. Similar to other reports [ 6 , 22 , 51 , 52 ], our results highlight how cell response to topography is highly dependent on intracellular context, even between cells of the same functional type.…”

Section: Resultssupporting

confidence: 91%

Customizable, engineered substrates for rapid screening of cellular cues

et al. 2020

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Biophysical cues robustly direct cell responses and are thus important tools for in vitro and translational biomedical applications. High throughput platforms exploring substrates with varying physical properties are therefore valuable. However, currently existing platforms are limited in throughput, the biomaterials used, the capability to segregate between different cues and the assessment of dynamic responses. Here we present a multiwell array (3 × 8) made of a substrate engineered to present topography or rigidity cues welded to a bottomless plate with a 96-well format. Both the patterns on the engineered substrate and the well plate format can be easily customized, permitting systematic and efficient screening of biophysical cues. To demonstrate the broad range of possible biophysical cues examinable, we designed and tested three multiwell arrays to influence cardiomyocyte, chondrocyte and osteoblast function. Using the multiwell array, we were able to measure different cell functionalities using analytical modalities such as live microscopy, qPCR and immunofluorescence. We observed that grooves (5 μ m in size) induced less variation in contractile function of cardiomyocytes. Compared to unpatterned plastic, nanopillars with 127 nm height, 100 nm diameter and 300 nm pitch enhanced matrix deposition, chondrogenic gene expression and chondrogenic maintenance. High aspect ratio pillars with an elastic shear modulus of 16 kPa mimicking the matrix found in early stages of bone development improved osteogenic gene expression compared to stiff plastic. We envisage that our bespoke multiwell array will accelerate the discovery of relevant biophysical cues through improved throughput and variety.

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Section: Resultssupporting

confidence: 91%

Customizable, engineered substrates for rapid screening of cellular cues

et al. 2020

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“…Any organization in which densely clustered Formins, that are elongating actin filaments in opposite orientations, could be sufficient to generate in-plane rotations. Consistent with this proposition are earlier reports in cultured cells showing that Formins anchored at peripheral adhesion sites, that are elongating actin filaments towards the cell center, are driving chiral in-plane swirling [25,27].…”

Section: Cyk-1/formin Activity In Active Rhoa Foci Promotes In-plane supporting

confidence: 86%

CYK-1/Formin activation in cortical RhoA signaling centers promotes organismal left-right symmetry breaking

Middelkoop

Garcia-Baucells

Quintero-Cadena

et al. 2021

Preprint

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1AbstractProper left-right symmetry breaking is essential for animal development and in many species the actin cytoskeleton plays an instrumental role in this process. Active torque generation in the actomyosin layer promotes left-right symmetry breaking in C. elegans embryos by driving chiral counter-rotating cortical flows. While both Formins and Myosins have been implied in left-right symmetry breaking, and both can rotate actin filaments in vitro, it remains unclear if active torques in the actomyosin cortex are generated by Formins, Myosins, or both. We combined the strength of C. elegans genetics with quantitative imaging and thin film, chiral active fluid theory to show that, while Non-Muscle Myosin II activity drives cortical actomyosin flows, it is permissive for chiral counter-rotation and dispensable for chiral symmetry breaking of cortical flows. Instead, we find that CYK-1/Formin activation in RhoA foci is instructive for chiral counter-rotation and promotes in-plane, active torque generation in the actomyosin cortex. Notably, we observe that artificially generated large active RhoA patches undergo rotations with consistent handedness in a CYK-1/Formin-dependent manner. Altogether, we conclude that, CYK-1/Formin-dependent active torque generation facilitates chiral symmetry breaking of actomyosin flows and drives organismal left-right symmetry breaking in the nematode worm.2SignificanceActive torque generation in the actin cytoskeleton has been implicated in driving left-right symmetry breaking of developing embryos, but which molecules generate the active torque and how active torque generation is organized subcellularly remains unclear. This study shows that cortical Formin, recruited to cortical regions where RhoA signaling is active, promotes active torque generation in the actomyosin layer. We find that active torque tends to locally rotate the cortex in a clockwise fashion, which drives the emergence of chiral counter-rotating flows with consistent handedness and facilitates left-right symmetry breaking of C. elegans embryos.

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“…Studies on animal cells have shown the impact of geometry on the organization of actin and MTs ( 14 , 16 , 18 , 68 , 69 ). When cells are allowed to spread on 2D patterns of various geometries, actin exhibits different organizations, with alignment along the long axis of elongated shapes or along the diagonal of square shapes ( 14 , 15 ).…”

Section: Conclusion/discussionmentioning

confidence: 99%

Cytoskeletal organization in isolated plant cells under geometry control

Durand-Smet

Spelman

Meyerowitz

et al. 2020

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

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The cytoskeleton plays a key role in establishing robust cell shape. In animals, it is well established that cell shape can also influence cytoskeletal organization. Cytoskeletal proteins are well conserved between animal and plant kingdoms; nevertheless, because plant cells exhibit major structural differences to animal cells, the question arises whether the plant cytoskeleton also responds to geometrical cues. Recent numerical simulations predicted that a geometry-based rule is sufficient to explain the microtubule (MT) organization observed in cells. Due to their high flexural rigidity and persistence length of the order of a few millimeters, MTs are rigid over cellular dimensions and are thus expected to align along their long axis if constrained in specific geometries. This hypothesis remains to be tested in cellulo. Here, we explore the relative contribution of geometry to the final organization of actin and MT cytoskeletons in single plant cells of Arabidopsis thaliana. We show that the cytoskeleton aligns with the long axis of the cells. We find that actin organization relies on MTs but not the opposite. We develop a model of self-organizing MTs in three dimensions, which predicts the importance of MT severing, which we confirm experimentally. This work is a first step toward assessing quantitatively how cellular geometry contributes to the control of cytoskeletal organization in living plant cells.

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Actin cytoskeleton self-organization in single epithelial cells and fibroblasts under isotropic confinement

Cited by 13 publications

References 44 publications

Customizable, engineered substrates for rapid screening of cellular cues

Customizable, engineered substrates for rapid screening of cellular cues

CYK-1/Formin activation in cortical RhoA signaling centers promotes organismal left-right symmetry breaking

Cytoskeletal organization in isolated plant cells under geometry control

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