A tethering complex drives the terminal stage of SNARE-dependent membrane fusion

Tethering proteins, known to mediate initial recognition and attachment during membrane fusion, are essential for driving the transition from the hemifused state to fusion pore formation. Tether proteins unleash SNARE Membrane fusion affects many cellular events and is essential for cell survival. It is thought that the various factors that mediate this process have distinct functions. For example, tethering proteins are responsible for the initial recognition and attachment of the fusing membranes, whereas SNARE protein complexes do the rest, providing mechanical energy to distort membranes through a hemifusion intermediate to form a fusion pore. But Andreas Mayer and colleagues now provide evidence that the division of labour is not so clear cut and that the tethering factors are not the lesser partner. They find that, in yeast cells, tether proteins are also essential for the transition from the hemifused state to the fusion pore. Specifically, these proteins play a mechanical part by increasing the volume of SNARE complexes and deforming the site of hemifusion. This lowers the energy barrier for pore opening. Membrane fusion in eukaryotic cells mediates the biogenesis of organelles, vesicular traffic between them, and exo- and endocytosis of important signalling molecules, such as hormones and neurotransmitters. Distinct tasks in intracellular membrane fusion have been assigned to conserved protein systems. Tethering proteins mediate the initial recognition and attachment of membranes, whereas SNARE (soluble N -ethylmaleimide-sensitive factor attachment protein receptor) protein complexes are considered as the core fusion engine. SNARE complexes provide mechanical energy to distort membranes and drive them through a hemifusion intermediate towards the formation of a fusion pore 1 , 2 , 3 . This last step is highly energy-demanding 4 , 5 . Here we combine the in vivo and in vitro fusion of yeast vacuoles with molecular simulations to show that tethering proteins are critical for overcoming the final energy barrier to fusion pore formation. SNAREs alone drive vacuoles only into the hemifused state. Tethering proteins greatly increase the volume of SNARE complexes and deform the site of hemifusion, which lowers the energy barrier for pore opening and provides the driving force. Thereby, tethering proteins assume a crucial mechanical role in the terminal stage of membrane fusion that is likely to be conserved at multiple steps of vesicular traffic. We ther