Research Projects

Integrin adhesion

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Actin in dendritic spines

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Super-resolution developments

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Integrin Molecular Dynamics: activation and conformation (Olivier Rossier)

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Expertise

Single protein tracking sptPALM

Super-resolution dSTORM, DNA-PAINT

Biophysical tools: Cell stretching

News

Deciphering mechano-biology using super-resolution microscopy

Cell stretching is amplified by active actin remodeling to deform and recruit proteins in mechano-sensitive structures Detection and conversion of mechanical forces into biochemical signals control cell functions during physiological and pathological processes. Mechano-sensing is based on protein deformations and reorganizations, yet the molecular mechanisms are still unclear. Using a cell stretching device compatible with super-resolution microscopy (SRM) and single protein tracking (SPT), we explored the nanoscale deformations and reorganizations of individual proteins inside mechano-sensitive structures. We achieved SRM after live stretching on intermediate filaments, microtubules and integrin adhesions. Simultaneous SPT and stretching showed that while integrins follow the elastic deformation of the substrate, actin filaments and talin also displayed lagged and transient inelastic responses associated with active acto-myosin remodeling and talin deformations. Capturing acute reorganizations of single-molecule during stretching showed that force-dependent vinculin recruitment is delayed and depends on the maturation of integrin adhesions. Thus, cells respond to external forces by amplifying transiently and locally cytoskeleton displacements enabling protein deformation and recruitment in mechano-sensitive structures.

Authors: Sophie Massou*, Filipe Nunes Vicente*, Franziska Wetzel*, Amine Mehidi, Dan Strehle, Cecile Leduc, Raphaël Voituriez, Olivier Rossier, Pierre, Nassoy and Gregory Giannone
* First co-authors

Nature Cell Biology, DOI 10.1038/s41556-020-0548-2.
Contacts IINS: Grégory Giannone and Filipe Nunes Vicente

+ Cf. INSB website (French) here
+ Cf. Bordeaux Neurocampus website here

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Orré T., Rossier O. and Giannone G. in Nature Communications - May 2021

Focal adhesions (FAs) initiate chemical and mechanical signals involved in cell polarity, migration, proliferation and differentiation. Super-resolution microscopy revealed that FAs are organized at the nanoscale into functional layers from the lower plasma membrane to the upper actin cytoskeleton. Yet, how FAs proteins are guided into specific nano-layers to promote interaction with given targets is unknown. Using single protein tracking, super-resolution microscopy and functional assays, we link the molecular behavior and 3D nanoscale localization of kindlin with its function in integrin activation inside FAs. We show that immobilization of integrins in FAs depends on interaction with kindlin. Unlike talin, kindlin displays free diffusion along the plasma membrane outside and inside FAs. We demonstrate that the kindlin Pleckstrin Homology domain promotes membrane diffusion and localization to the membrane-proximal integrin nano-layer, necessary for kindlin enrichment and function in FAs. Using kindlin-deficient cells, we show that kindlin membrane localization and diffusion are crucial for integrin activation, cell spreading and FAs formation. Thus, kindlin uses a different route than talin to reach and activate integrins, providing a possible molecular basis for their complementarity during integrin activation.

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Mechanical control of actin regulators during cell migration - Nature Cell Biology, Nov. 2021

Actin filaments generate mechanical forces that drive membrane movements during trafficking, endocytosis and cell migration. Reciprocally, adaptations of actin networks to forces regulate their assembly and architecture. Yet, a demonstration of forces acting on actin regulators at actin assembly sites in cells is missing. Here we show that local forces arising from actin filament elongation mechanically control WAVE regulatory complex (WRC) dynamics and function, that is, Arp2/3 complex activation in the lamellipodium. Single-protein tracking revealed WRC lateral movements along the lamellipodium tip, driven by elongation of actin filaments and correlating with WRC turnover. The use of optical tweezers to mechanically manipulate functional WRC showed that piconewton forces, as generated by single-filament elongation, dissociated WRC from the lamellipodium tip. WRC activation correlated with its trapping, dwell time and the binding strength at the lamellipodium tip. WRC crosslinking, hindering its mechanical dissociation, increased WRC dwell time and Arp2/3-dependent membrane protrusion. Thus, forces generated by individual actin filaments on their regulators can mechanically tune their turnover and hence activity during cell migration.

Authors: Amine Mehidi, Frieda Kage, Zeynep Karatas, Maureen Cercy, Matthias Schaks, Anna Polesskaya, Matthieu Sainlos, Alexis Gautreau, Olivier Rossier, Klemens Rottner and Grégory Giannone

Forces generated by lamellipodial actin filament elongation regulate the WAVE complex during cell migration
Nature Cell Biology, 23, pages 1148–1162 (2021). https://www.nature.com/articles/s41556-021-00786-8

*Contact IINS: Grégory Giannone

+ Cf. INSB website (French) here
+ Cf. Bordeaux Neurocampus website here

Gregory Gianonne and his team win the FRM Team Price 2023

Created in 1948, the Fondation pour la Recherche Medicale (FRM) has as an aim to support and fund the public research in every medical and pathophysiologic fields.

Thus, the FRM supports more than 400 new researches conducted in the laboratories of public research and higher education organisations every year (INSERM, CNRS, INRA, CEA, Universities, Prestigious Universities, health institution, …).

Gregory Giannone (CNRS researcher and team leader “Mechano-biology of motile and neuronal structure”) and his team have just won the FRM Team Price 2023!                                                      

This price is awarded to teams proposing an innovative research program in biology with potential health applications.

Gregory Giannone’s team explains us their project:

"Adhesive and cytoskeletal structures control cell functions such as migration and proliferation. As such they regulate cell behavior during physiological processes such as development; but when altered, these structures contribute to pathologies including cancer. How assembly of adhesive and cytoskeletal structures and resulting mechanical forces are coordinated to shape cell movements and morphologies during development and cancer progression remains fundamental questions. Despite recent advances in imaging methods in multicellular environments (organoids, small organisms), a molecular understanding of these fundamental processes is still lacking. To reach this molecular understanding, we developed for the last ten years, new strategies to study integrin adhesions and actin-based protrusions at the molecular level using super-resolution microscopy and single protein tracking. We unraveled key molecular events leading to: integrins activation and mechano-sensing in healthy and cancer cells; actin assembly in dendritic spines, and in lamellipodia. However, these findings were obtained by studying isolated cells on stiff 2D substrates. In this proposal, we aim to reach a molecular understanding of cell movements and morphologies in 3D multicellular assemblies characterized by softer, confined and dynamic 3D environments. In this project, we will decrypt the mechanical and biochemical molecular rules that govern the assembly, dynamics and coordination of integrin adhesions and actin protrusions in 3D multicellular systems during two fundamental processes: (AIM 1) the formation of long-lasting macromolecular complexes in vivo during development of integrin-based muscle attachment sites in Drosophila; (AIM 2) the formation of transient macromolecular complexes supporting the ability of Small Cell Lung Cancer cells to assemble into spheroids and to migrate during metastasis. This project could help to find new and more specific therapeutic strategies against this cancer."

Congratulations to the "Mechano-biology of motile and neuronal structure" team!

Selected Publications

Nunes Vicente F., Rossier O., Giannone G.

Super-resolution microscopy: A window into mechanobiology at the molecular level Medecine sciences : M/S (2022)
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Nunes Vicente F., Chen T., Rossier O., Giannone G.

Novel imaging methods and force probes for molecular mechanobiology of cytoskeleton and adhesion Trends in Cell Biology (2022)
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Orré T., Joly A., Karatas Z., Kastberger B., Cabriel C., Böttcher R. T., Lévêque-Fort S., Sibarita JB., Fässler R., Wehrle-Haller B., O. Rossier*, G. Giannone*

Molecular motion and tridimensional nanoscale localization of kindlin control integrin activation in focal adhesions Nature Communications (2021)
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Souissi M., Pernier J., Rossier O., Giannone G., Le Clainche C., Helfer E., Sengupta K.

Integrin-Functionalised Giant Unilamellar Vesicles via Gel-Assisted Formation: Good Practices and Pitfalls. International journal of molecular sciences (2021)
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Mehidi A., Kage F., Karatas Z., Cercy M., Schaks M., Polesskaya A., Sainlos M., Gautreau A., Rossier O., Rottner K. and Giannone G.

Forces generated by lamellipodial actin filament elongation regulate the WAVE complex during cell migration. Nature cell biology (2021)
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Massou S., Nunes Vicente F., Wetzel F., Mehidi A., Strehle D., Leduc C., Voituriez R., Rossier O., Nassoy P., Giannone G.

Cell stretching is amplified by active actin remodelling to deform and recruit proteins in mechanosensitive structures Nature Cell Biology (2020)
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Mehidi A., Rossier O., Schaks M., Chazeau A., Biname F., Remorino A., Coppey M., Karatas Z., Sibarita J.B., Rottner K., Moreau V., Giannone G.

Transient Activations of Rac1 at the Lamellipodium Tip Trigger Membrane Protrusion Current Biology (2019)
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Paszek M.J., Dufort C.C., Rossier O., Bainer R., Mouw J.K., Godula K., Hudak J.E., Lakins J.N., Wijekoon A., Cassereau L., Rubashkin M.G., Magbanua M.J., Thorn K.S., Davidson M.W., Rugo H.S., Park J.W., Hammer D.A, Giannone G., Bertozzi C.R, Weaver V.M.

The cancer cell glycocalyx mechanically primes integrin-dependent growth and survival Nature (2014)
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Chazeau A., Mehidi A., Nair D., Gautier J., Leduc C., Chamma I., Kage F., Kechkar A., Thoumine O., Rottner K., Choquet D., Gautreau A., Sibarita J.B., Giannone G.

Nanoscale segregation of actin nucleation and elongation factors determines dendritic spine protrusion. EMBO Journal (2014)
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Rossier O., Octeau V., Sibarita J.B., Leduc C., Tessier B., Nair D., Gatterdam V., Destaing O., Albiges-Rizo C., Tampé R., Cognet L, Choquet D., Lounis B., and Giannone G.

Integrins β1 and β3 exhibit distinct dynamic nanoscale organizations inside focal adhesions. Nature Cell Biology (2012)
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Giannone G., Hosy E., Levet F., Constals A., Schulze K., Sobolevsky A. I., Rosconi M. P., Gouaux E., Tampé R., Choquet D. and Cognet L.

Dynamic super-resolution imaging of endogenous proteins on living cells at ultra-high density. Biophysical Journal (2010)
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