Effet of acto-myosin dynamics on membrane nanotube stability – DynAcTube

To address these questions, I will perform in vitro experiments to decipher the physics of nanotube remodeling in biochemically controlled assays recapitulating key aspects of cellular membranes and actin dynamics. I will use two complementary techniques to form membrane nanotubes on which I will reconstitute acto-myosin networks from purified proteins. By using optical tweezers, I will measure the forces implied in nanotube formation and maintenance in presence of acto-myosin. In parallel, I develop a novel assay to image supported nanotubes and acto-myosin networks polymerizing on such nanotubes at the nanometric scale by using Atomic Force Microscopy. By combining these two techniques, I will show how the structure of the acto-myosin network at the nanometric scale dictates tube reshaping at the micrometric scale and how this explains the results obtained in cells. This will shed new light on nanotube shape regulation and deepen our understanding of cellular functions.

In Lamour et al., Phys. Rev. X, 2020, we propose a nanomechnical mapping of membrane nanotubes by AFM. We relate their morphology and their mechanical properties and show how they are affected in presence of an actin coat on the tube surface.

In Allard et al., Sci. Adv. 2020, we polymerize a dynamic actin network at the surface of a nanotube and follow how actin affect its fate when it is further pulled, as is it the case in vivo. Depending on the actin amount on the tube, it gets completely stabilized or can be elongated. In this latter case, the actin coat tears under elongation and the tube becomes transiently heterogeneous in radius: newly formed regions devoid of actin are smaller and could be available for other proteins that remodel tubes.

By fulfilling the tasks detailed above, I will obtain crucial and novel information on the effect of acto-myosin dynamics on membrane nanotube remodeling for various acto-myosin network and membrane compositions. This interdisciplinary biophysical approach will contribute to the understanding of phenomena such as intracellular traffic and endocytosis that are fundamental processes conserved in all eukaryotic cells. I will also develop a novel assay allowing accessing the nanostructure of membrane nanotubes by AFM.

1. Allard, A.; Valentino, F.; Sykes, C.; Betz, T.; Campillo, C.* «Fluctuations of a membrane nanotube interacting with an actin network», Phys. Rev. E, accepted
2. Allard, A.; Bouzid, M.; Betz, T.; Simon, C.; Abou-Ghali, M.; Lemière, J.; Valentino, F.; Manzi, J.; Brochard-Wyart, F.; Guevorkian, K.; Plastino, J.; Lenz, M.; Campillo, C.*; Sykes, C.* «Effect of an actin sleeve on a membrane nanotube», Science Advances, 6 (17), eeaz 3050, 2020
3. Lamour, G.; Allard, A.; Pelta, J.; Labdi, S.; Lenz, M.; Campillo, C.* « Morphology and mechanical properties of membrane nanotubes by atomic force microscopy », Phys Rev X, 10, 011031, 2020

Inside living cells, the remodeling of membrane nanotubes by actomyosin networks is crucial for processes such as intracellular traffic or endocytosis. However, the mechanisms by which actomyosin dynamics affect nanotube morphology are largely unknown. How much radial and axial forces are generated on a nanotube by actomyosin dynamics? Can these forces lead to nanotube scission? How do they relate to the structure of the actomyosin network? To address these questions, I will perform in vitro experiments to decipher the physics of nanotube remodeling in biochemically-controlled assays recapitulating key aspects of cellular membranes and actin dynamics. I will two complementary techniques to form membrane nanotubes on which I reconstitute actomyosin networks from purified proteins. By using optical tweezers, I will measure the forces implied in nanotube formation and maintenance in presence of actomyosin. In parallel, I develop a novel assay to image nanotubes and actomyosin networks polymerizing these nanotubes by Atomic Force Microscopy. By combining these two techniques, I will investigate how the structure of the actomyosin network at the nanometric scale dictates tube reshaping at the micrometric scale and how this explains the tube remodeling observed in cells. This will shed new light on nanotube shape regulation and deepen our understanding of cellular functions. I have been working on membrane nanotubes and reconstituted actin networks for over ten years and I have used AFM for over four years, therefore I have the skills and the best environment, in my lab and with my collaborators, to develop this novel and ambitious project The JCJC grant will allow me to have at least two collaborators under my supervision, obtain important scientific results and thus to strengthen my scientific independence and foster my career development.