Extended Synaptotagmin is a presynaptic ER Ca²⁺ sensor that promotes neurotransmission and synaptic growth in Drosophila

ABSTRACT

The endoplasmic reticulum (ER) is an extensive presynaptic organelle, exerting important influences at synapses by responding to Ca²⁺ and modulating transmission, growth, lipid metabolism, and membrane trafficking. Despite intriguing evidence for these crucial functions, how presynaptic ER influences synaptic physiology remains enigmatic. To gain insight into this question, we have generated and characterized mutations in the single Extended Synaptotagmin (Esyt) ortholog in Drosophila. Esyts are evolutionarily conserved ER proteins with Ca²⁺ sensing domains that have recently been shown to orchestrate membrane tethering and lipid exchange between the ER and plasma membrane. We first demonstrate that Esyt localizes to an extensive ER structure that invades presynaptic terminals at the neuromuscular junction. Next, we show that synaptic growth, structure, function, and plasticity are surprisingly unperturbed at synapses lacking Esyt expression. However, presynaptic overexpression of Esyt leads to enhanced synaptic growth, neurotransmission, and sustainment of the vesicle pool during intense levels of activity, suggesting that elevated Esyt at the ER promotes constitutive membrane trafficking or lipid exchange with the plasma membrane. Finally, we find that Esyt mutants fail to maintain basal neurotransmission and short term plasticity at elevated extracellular Ca²⁺, consistent with Esyt functioning as an ER Ca²⁺ sensor that modulates synaptic activity. Thus, we identify Esyt as a presynaptic ER Ca²⁺ sensor that can promote neurotransmission and synaptic growth, revealing the first in vivo neuronal functions of this conserved gene family.