The synaptotagmin juxtamembrane domain is involved in neuroexocytosis

• The highly cationic juxtamembrane segment of synaptotagmin juxtamembrane domain was synthesized.
• This peptide inhibits neurotransmitter release at the neuromuscular junction of mice and Drosophila.
• This peptide localizes mainly on the presynaptic membrane.
• The synaptotagmin juxtamembrane peptide binds monophosphoinositides in a Ca2+-independent manner.
• The juxtamembrane segment of synaptotagmin may contribute to the formation of the hemifusion intermediate.

Synaptotagmin is a synaptic vesicle membrane protein which changes conformation upon Ca2+ binding and triggers the fast neuroexocytosis that takes place at synapses. We have synthesized a series of peptides corresponding to the sequence of the cytosolic juxtamembrane domain of synaptotagmin, which is highly conserved among different isoforms and animal species, with or without either a hexyl hydrophobic chain or the hexyl group plus a fluorescein moiety. We show that these peptides inhibit neurotransmitter release, that they localize on the presynaptic membrane of the motor axon terminal at the neuromuscular junction and that they bind monophosphoinositides in a Ca2+-independent manner. Based on these findings, we propose that the juxtamembrane cytosolic domain of synaptotagmin binds the cytosolic layer of the presynaptic membrane at rest. This binding brings synaptic vesicles and plasma membrane in a very close apposition, favouring the formation of hemifusion intermediates that enable rapid vesicle fusion.