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. 2011 May 31;108(22):9107-12.
doi: 10.1073/pnas.1018652108. Epub 2011 May 13.

Integrin adhesion drives the emergent polarization of active cytoskeletal stresses to pattern cell delamination

C Meghana  1 , Nisha Ramdas, Feroz Meeran Hameed, Madan Rao, G V Shivashankar, Maithreyi Narasimha
Affiliations

Integrin adhesion drives the emergent polarization of active cytoskeletal stresses to pattern cell delamination

C Meghana et al. Proc Natl Acad Sci U S A. .

Abstract

Tissue patterning relies on cellular reorganization through the interplay between signaling pathways and mechanical stresses. Their integration and spatiotemporal coordination remain poorly understood. Here we investigate the mechanisms driving the dynamics of cell delamination, diversely deployed to extrude dead cells or specify distinct cell fates. We show that a local mechanical stimulus (subcellular laser perturbation) releases cellular prestress and triggers cell delamination in the amnioserosa during Drosophila dorsal closure, which, like spontaneous delamination, results in the rearrangement of nearest neighbors around the delaminating cell into a rosette. We demonstrate that a sequence of "emergent cytoskeletal polarities" in the nearest neighbors (directed myosin flows, lamellipodial growth, polarized actomyosin collars, microtubule asters), triggered by the mechanical stimulus and dependent on integrin adhesion, generate active stresses that drive delamination. We interpret these patterns in the language of active gels as asters formed by active force dipoles involving surface and body stresses generated by each cell and liken delamination to mechanical yielding that ensues when these stresses exceed a threshold. We suggest that differential contributions of adhesion, cytoskeletal, and external stresses must underlie differences in spatial pattern.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Cellular dynamics associated with subcellular cytoplasmic laser perturbation. (A) Stage 14 embryo showing the contour of the AS highlighting a cohort of cells with the delaminating cell (DC, purple) surrounded by its nearest neighbours (lilac). Schematic representation of the adhesive interfaces that enable cellular interactions in the AS (B) and cytoskeletal organization at the LE–pAS cell interface (C). (D1–D5) Time-lapse confocal images of an embryo labeled with E-Cadherin GFP. (E) Representative graph of apical surface area dynamics showing typical velocities (mean ± SEM) of constriction (n = 5). (E′) Frequency distribution of prestress. (F) Fractional change in apical surface area of DC (black), its NN (dark gray, n = 5), and DN (light gray, n = 4). (G) Strain anisotropy of NN (colored) and apical surface area of the DC (black). (H) Centroid position of the NN with respect to the DC (n = 9). Asterisks indicate perturbed cell. Analysis of fractional change in area and perimeter, strain anisotropy, and circularity changes of multiple cohorts can be found in Fig. S1. (Scale bar, 10 μm.)
Fig. 2.
Fig. 2.
Emergent polarization of the cellular cytoskeleton in response to subcellular laser perturbation. Time-lapse confocal images of sqhGFP (A1–A8), actin5CGFP (B), and EB1GFP (C) to show actin, myosin, and microtubule dynamics upon perturbation. Arrows point to particulate streams of sqh (A1–A4), enrichment at the DC–NN interface (A5–A8), actin ruffles in the neighboring cells (B; the ablated cell is actin GFP negative), and polarized microtubule reorganization (C). Apical projections and orthogonal (XZ) views are shown. Asterisks indicate the ablated cell. See Fig. S5 for response to perturbation of tubulin GFP and microtubule organization in fixed ablated embryos. (Scale bars, 10 μm.)
Fig. 3.
Fig. 3.
Adhesion-dependent mechanical compliance and cytoskeletal polarization. Time-lapse confocal images of an embryo labeled with E-Cadherin GFP showing anisotropic expansion (A1, arrow; n = 5) and aspect ratio dynamics of pAS cells upon perturbation (A2). Dynamics of fractional change in apical surface area induced by perturbation in control cells (black in B and C) and upon expression (gray) of αPS3 RNAi (B; n = 8) or spastin (C; n = 8). Time-lapse confocal images showing response of actin (D and E) and myosin (F–H) to cytoplasmic perturbation in control (D and F), βPS RNAi (E and H), and spastin-overexpressing embryos (G). Asterisks mark perturbed cell. Response of actin to perturbation in αPS3 RNAi can be found in Fig. S6. (Scale bars, 10 μm.)
Fig. 4.
Fig. 4.
Temporal hierarchies of molecular and physical events during natural and induced delamination. Schematic representations of cell area dynamics, shape transformations, cytoskeletal dynamics, and cell behaviors that accompany induced (B, dark gray arrows) and natural delamination (B, light gray arrows) in wild-type embryos are depicted along the timeline beginning at t0, marking the onset of induction, and te, marking the time of cell extrusion; delaminating cell is at the center of the rosette in all images. An integrin-deficient rosette is shown below it. The times of onset from t0 at which the regimes that characterize the dynamics (A, Right) become evident are expressed as means computed from multiple movies (numbers in brackets are SEs of mean; n = 8 for actin, 6 for microtubules, and 5 for myosin) and their relation to changes in area that accompany induced delamination (A, Left). A0 and Amax are initial and maximally expanded normalized areas.
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