> Retour à la liste des séminaires


Mardi 19 Novembre 2019
De 14h à 15h
Centre de recherche - Paris - Amphithéâtre Antoine Lacassagne

“Dynamics of membrane tension, synaptic vesicle recycling, and modulation of fusion pores by the neuronal calcium sensor Synaptotagmin-1”

Calcium-triggered release of neurotransmitters and hormones entails fusion of cargo-filled vesicles with the plasma membrane (exocytosis). Although key molecules involved in this process and their basic functions have been identified, how these components work together to achieve release well within a millisecond after stimulation is not understood. After fusion, plasma membrane area returns to its pre-release value via endocytosis over much slower time scales (0.1-10 seconds). Two key factors that regulate release and vesicle recycling are dynamics of the initial nanometer-sized fusion pore and membrane flows.

During release, some fusion pores fluctuate in size, flicker open-closed multiple times, and either reseal or dilate. Pore resealing results in recovery of a nearly intact vesicle-- a rapid mechanism for vesicle recycling. To monitor dynamics of single fusion pores, we fuse ~25 nanometer phospholipid bilayer discs (nanodiscs) with engineered cells expressing components of the neuronal fusion machinery on their surfaces. The nanodiscs are reconstituted with complementary fusion proteins. Fusion results in a nanometer-sized pore whose dynamics are monitored using cell-attached, voltage-clamp recordings of currents passing through the pore. The method allows probing single pores with sub-millisecond time resolution. I will present how molecular crowding and mechanical forces generated by Synaptotagmin-SNARE-membrane interactions lead to larger pores.

Another crucial factor regulating synaptic transmission by neurons and hormone release by neuroendocrine cells is membrane tension. In neurons, endocytosis occurs some distance away from the active zone where release takes place. Because exocytosis and endocytosis locally decrease and increase membrane tension, respectively, sustained exo-endocytic activity would lead to membrane tension gradients. Such gradients relax by membrane flows, which are extremely slow in plasma membranes of non-neuronal cells, or non-terminal regions of neurons. Slow membrane flows could limit spatio-temporal coupling of exo- and endocytosis, but have never been probed in presynaptic terminals. We characterized membrane tension dynamics in presynaptic terminals by pulling thin membrane tethers from them. We find extremely facile membrane flows and tension equilibration at presynaptic plasma membranes, which appear to be tuned for rapid turnover of synaptic vesicles, thus playing a key role in neurotransmission. 


Erdem Karatekin

Nanobiology Institute, Yale University, New Haven

Invité(e)(s) par

Françoise Brochard