Cell migration over heterogeneous substrates during wound healing or morphogenetic processes leads to shape changes driven by different organizations of the actin cytoskeleton and by functional changes including lamellipodial protrusions and contractile actin cables. Cells distinguish between cell-sized positive and negative curvatures in their physical environment by forming protrusions at positive curvatures and actin cables at negative curvatures; however, the cellular mechanisms remain unclear. Here, we report that concave edges promote polarized actin structures with actin flow directed towards the cell edge, in contrast to well-documented retrograde flow at convex edges. Anterograde flow and contractility induce a tension anisotropy gradient. A polarized actin network is formed, accompanied by a local polymerization–depolymerization gradient, together with leading-edge contractile actin cables in the front. These cables extend onto non-adherent regions while still maintaining contact with the substrate through focal adhesions. The contraction and dynamic reorganization of this actin structure allows forward movements enabling cell migration over non-adherent regions on the substrate. These versatile functional structures may help cells sense and navigate their environment by adapting to external geometric and mechanical cues.
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The authors thank D. Delacour, N. Gov, R.-M. Mège, group members from MBI and IJM for helpful discussions and critical reading of the manuscript. The authors would also like to thank MBI Microfabrication (G. Grenci and M. Asraf), MBI Science Communication Core (C.X. Wong and S. Wolf) and MBI Microscopy Core (F. Margadant) for continuous support. The authors are grateful for the ImagoSeine core facility of the Institut Jacques Monod, member of IBiSA and the France-BioImaging (ANR-10-INBS-04) infrastructure for their support. The authors thank A. Bershadsky, M. Sheetz and P. Kanchanawong for sharing of plasmid constructs. The authors are grateful to W. J. Nelson and F. Martin-Belmonte for their generous gift of MDCK cell lines. Financial support from the Mechanobiology Institute, the European Research Council under the European Union’s Seventh Framework Programme (FP7/2007-2013) / ERC grant agreement numbers 617233 (B.L.) and 616480 (X.T.), USPC-NUS collaborative program, Agence Nationale de la Recherche (ANR) (“POLCAM” (ANR-17- CE13-0013), CODECIDE (ANR-17-CE-13-0022)), the LABEX ‘Who am I?’, and the Spanish Ministry of Economy and Competitiveness (BFU2015-65074-P) are gratefully acknowledged. T.C. acknowledges the USPC-NUS international PhD mobility programme. T.S. is supported by the Israel Science Foundation (grant number 921/15).
The authors declare no competing interests
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Supplementary Sections 1–4, Supplementary References 1–10, Supplementary Figures 1–6, Supplementary Table 1, Supplementary Video Captions 1–16.
MDCK cells migrate over a non-adherent circular island. Images were taken with wide field TIRF, mApple Actin shown in magenta and GFP MRLC shown in green. Scale bar, 10 µm.
Phase contrast video showing MDCK WT cells on fibronectin pattern (magenta line). Scale bar, 10 µm.
Live SIM video of MDCK GFP actin cell at negative curvature on fibronectin pattern (magenta line) shows anterograde flow. See Fig. 3a for colour code. Scale bar, 5 µm.
Photoconversion of radial fibre in mEos-actin (before conversion-green, after conversion-magenta) transfected MDCK cell at negative curvature on fibronectin pattern (white line). Images were acquired with a scanning confocal microscope showing anterograde flow of radial fibres. Scale bar, 2 µm.
Fluorescent speckle microscope of mEmerald-actin transfected MDCK cell at negative curvature on fibronectin pattern (magenta line) shows anterograde flow of speckles. Imaged with a wide-field TIRF microscope. Scale bar, 5 µm.
Fluorescent speckle microscope of mEmerald-actin transfected MDCK cell at positive curvature on fibronectin pattern (magenta line) shows retrograde flow of speckles. Imaged with a wide-field TIRF microscope. Scale bar, 5 µm.
Laser ablations of TFs in a MDCK cell expressing GFP-MRLC at negative curvature imaged with a scanning confocal microscope. Scale bar, 5 µm.
Myosin flow in control, blebbistatin treated and washout cells. MCDK cells were transfected with mApple Myosin IIA and imaged with a spinning disk confocal microscope with a super-resolution module. Video shows the same cell imaged under the three conditions. Scale bar, 5µm.
Photoconversion of a square ROI region (yellow box) in a MDCK cell transfected with mEos2-actin at negative curvature (white line shows edge of fibronectin pattern), imaged with scanning confocal microscope. Green channel shows unconverted fluorescence, magenta channel shows converted fluorescence. Scale bar, 5 µm.
Live SIM video of MDCK GFP actin cells showing actin dynamics in the centre region at negative curvature. Cells were treated with 200 µM CK666 for 30 min before washout to remove the underlying actin fibres (due to contractility induced anterograde flow of previously formed fibres and no new fibre formation with inhibition of actin polymerization) and reveal the dynamics of newly polymerized actin filaments. Edge of fibronectin pattern is shown with a magenta line. Scale bar, 5 µm.
1 nM jasplakinolide treatment induced the collapse of actin structure. MDCK cells were transfected with mApple Actin (magenta) and GFP Myosin IIA (green) and imaged with a spinning disk confocal microscope with a super-resolution module. Edge of fibronectin pattern is shown with a white line. Scale bar, 5µm.
Severance of the radial fibres with laser ablation induces recruitment of Arp2/3 and polymerization of actin at the rear of the ablation site. Scanning confocal images are shown for mApple actin (left) and mEmerald Arp-p34 transfected MDCK cell (right). See Fig. 5k for colour code. White line marks the edge of fibronectin pattern. Scale bar, 5 µm.
Laser ablation of actin cable out of the adherent pattern (left) induced a retraction of the actin structure whereas ablation of actin cable on the adherent pattern (right) had no effect. MDCK GFP MRLC cells were imaged with a scanning confocal microscope. Yellow line marks the edge of the fibronectin pattern (30 µm radius negative curvature). Scale bar, 10 µm.
MDCK monolayer could not migrate over non-adherent circle of 100 µm diameter. MDCK GFP MRLC (green) MDCK cells were transfected with mApple Actin (magenta) and were imaged with a wide-field TIRF microscope. Fibronectin pattern is shown in cyan. Scale bar, 20 µm.
MDCK α-catenin KD cells migrate over non-adherent circle of 30 µm diameter. Cells were transfected with mApple Actin (magenta) and mEmerald Myosin IIA (green) and were imaged with a wide-field TIRF microscope. Fibronectin pattern is shown in cyan. Scale bar, 10 µm.
Laser ablation of actin cable at the edge of an expanding MDCK monolayer on a continuous substrate resulted in transformation into lamellipodium when the edge was of positive curvature (left) and resulted in actin cable repair when the edge was of negative curvature (right). Cells were transfected with mApple Actin and imaged with a scanning confocal microscope. Scale bar, 10 µm.
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Chen, T., Callan-Jones, A., Fedorov, E. et al. Large-scale curvature sensing by directional actin flow drives cellular migration mode switching. Nat. Phys. 15, 393–402 (2019). https://doi.org/10.1038/s41567-018-0383-6
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