Dynamin - Mediated Membrane Fission

The eucaryotic cell is organized in several compartments named organelles. These organelles and the cell itself are delimited by lipid membranes. The ssion of these membranes is required for the vesicular tra c between organelles. Endocytosis is the mechanism of vesicular tra c from the plasma membrane towards other organelles inside the cell. Dynamin is a guanosine triphosphatase implicated in vesicle scission during Clathrin-mediated endocytosis. It polymerizes into a helix at the neck of endocytic buds. Upon guanosine triphosphate hydrolysis (GTP), conformational changes modify the helical structure: the inner radius of the Dynamincoated tube decreases from 10 to 5 nm and the helical pitch reduces from 13 to 9 nm. These modi cations show that ssion proceeds through a constriction mechanism. The dynamics of Dynamin constriction is investigated by attaching microbeads along Dynamin-coated tubes and by monitoring the beads' rotations after GTP addition. I found that the deformation of Dynamin helices is highly concerted and damped by the friction between membrane and Dynamin. However constriction is not enough to trigger ssion. To further understand the ssion reaction, Dynamin polymerization and ssion are studied on lipid nanotubes extruded from Giant Unilamellar Vesicles. My work shows that ssion occurs at the edge of the Dynamin helix, where the membrane is strongly curved. A statistical analysis of ssion time reveals that the ssion reaction can be modelled by a single step rate-limiting energy barrier. The ssion time dependence on membrane tension, membrane rigidity and torque is established theoretically and validated experimentally. Dynamin torque is evaluated between 730 and 1100 pN.nm. The relevance of the tension dependence is con rmed in vivo. This work allows to give a quantitative picture of the energy landscape of Dynamin-mediated ssion: the height of the energy barrier of ssion is estimated around 70 kBT . te l-0 07 66 69 4, v er si on 1 18 D ec 2 01 2