During wound repair, branching morphogenesis and carcinoma dissemination, cellular rearrangements are fostered by a solid-to-liquid transition, known as unjamming. The biomolecular machinery behind unjamming and its pathophysiological relevance remain, however, unclear. Here, we study unjamming in a variety of normal and tumorigenic epithelial two-dimensional (2D) and 3D collectives. Biologically, the increased level of the small GTPase RAB5A sparks unjamming by promoting non-clathrin-dependent internalization of epidermal growth factor receptor that leads to hyperactivation of the kinase ERK1/2 and phosphorylation of the actin nucleator WAVE2. This cascade triggers collective motility effects with striking biophysical consequences. Specifically, unjamming in tumour spheroids is accompanied by persistent and coordinated rotations that progressively remodel the extracellular matrix, while simultaneously fluidizing cells at the periphery. This concurrent action results in collective invasion, supporting the concept that the endo-ERK1/2 pathway is a physicochemical switch to initiate collective invasion and dissemination of otherwise jammed carcinoma.
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The authors declare that all data supporting the findings of this study are available within the paper and its supplementary Information files and from the corresponding authors upon reasonable request.
The codes used for the analysis are all indicated in the Methods.
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This work has been supported by: the Associazione Italiana per la Ricerca sul Cancro (AIRC) to G.S. (IG#18621), P.P.D.F (IG#18988 and MCO 10.000), and F.G. (MFAG#22083); the Italian Ministry of University and Scientific Research (MIUR) to P.P.D.F. and G.S. (PRIN: PROGETTI DI RICERCA DI RILEVANTE INTERESSE NAZIONALE – Bando 2017#2017HWTP2K); the Italian Ministry of Health (RF-2013-02358446) to G.S. Regione Lombardia and CARIPLO foundation (Project 2016-0998) to R.C.; Worldwide Cancer Research (WCR#16-1245) to S.S. C.M. and F.G. are partially supported by fellowships from the University of Milan, E.B. from the FIRC-AIRC. We thank J. Christian (Max Planck Institute for Medical Research, Heidelberg, Germany) for help with fluorescent beads.
The authors declare no competing interests.
Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Figs. 1–12, Supplementary Tables 1–4, Supplementary Video Legends 1–29, Supplementary Discussion and Supplementary references 1–19
Supplementary Video 1
RAB5A reawakening of collective motion in jammed epithelial monolayers depends on EGF
Supplementary Video 2
RAB5A reawakening of collective motion in jammed epithelial monolayers depends on EGFR
Supplementary Video 3
EGF-dependence of RAB5A flocking motility in EGFP-H2B-jammed epithelia monolayers
Supplementary Video 4
PIV analysis of EGF and EGFR-dependent endocytic unjamming
RAB5A flocking motion in jammed epithelia is reduced by silencing Dynamin 2
Supplementary Video 6
RAB5A flocking motion in jammed epithelia is reduced by silencing RTN3, but not RTN4
Supplementary Video 7
RAB5A, but neither RAB5B nor RAB5C induces flocking motion in jammed epithelia
Supplementary Video 8
Flocking motion is abrogated by treatment with inhibitors of the MAPK/ERK1/2 pathway
Supplementary Video 9
Flocking motion is abrogated by treatment with Dynasore
Supplementary Video 10
MEK-DD is not sufficient to reawaken collective motion in jammed MCF10A monolayers
Supplementary Video 11
RAB5A-induced cryptic lamellipodia is compromised by treatment with MEK1/2 inhibitor
Supplementary Video 12
RAB5A-induced cryptic lamellipodia are inhibited by silencing the WAVE complex
Supplementary Video 13
RAB5A reawakening of collective motion in jammed epithelia is impaired by silencing NAP1
Supplementary Video 14
Silencing of NAP1 or WAVE2 affects RAB5A-induced wound closure in epithelial monolayers
Supplementary Video 15
Acini kinematic motility assay
Supplementary Video 16
PIV analysis on acini motility
Supplementary Video 17
RAB5A-mediated unjamming in MCF10A acini is trafficking-, EGFR- and ERK1/2-dependent
Supplementary Video 18
RAB5A-induced angular motion of MCF10A is independent of cell proliferation
Supplementary Video 19
RAB5A overcomes kinetic arrest of differentiated MCF10A acini
Supplementary Video 20
RAB5A reawakens collective motion in jammed MCF10.DCIS.com carcinoma cells
Supplementary Video 21
RAB5A promotes wound closure and flocking motion
Supplementary Video 22
RAB5A-mediated unjamming induces coordinated angular rotation in breast cancer spheroids
Supplementary Video 23
3D DVA of a RAB5A rotating spheroid after removal of the global rotation
Supplementary Video 24
RAB5A-flocking in spheroids is trafficking-, EGFR-, ERK1/2- and ARP2/3-dependent
Supplementary Video 25
RAB5A-mediated 3D unjamming promotes collective invasion in tumour spheroids
Supplementary Video 26
RAB5A spheroids exert larger stresses on surrounding ECM
Supplementary Video 27
Instantaneous velocity and stress maps of control and RAB5A-expressing spheroids.
Supplementary Video 28
RAB5A-mediated flocking promotes collective invasion in ex vivo DCIS tumour slices
Supplementary Video 29
PIV analysis on ex vivo DCIS tumour slice motility
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Palamidessi, A., Malinverno, C., Frittoli, E. et al. Unjamming overcomes kinetic and proliferation arrest in terminally differentiated cells and promotes collective motility of carcinoma. Nat. Mater. 18, 1252–1263 (2019). https://doi.org/10.1038/s41563-019-0425-1
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