Unc13A dynamically stabilizes vesicle priming at synaptic release sites for short-term facilitation and homeostatic potentiation

Presynaptic plasticity adjusts neurotransmitter (NT) liberation. Short-term facilitation (STF) tunes synapses to millisecond repetitive activation, while presynaptic homeostatic potentiation (PHP) of NT release stabilizes transmission over minutes. Despite different timescales of STF and PHP, our analysis of Drosophila neuromuscular junctions reveals functional overlap and shared molecular dependence on the release-site protein Unc13A. Mutating Unc13A’s calmodulin binding domain (CaM-domain) increases baseline transmission while blocking STF and PHP. Mathematical modeling suggests that Ca2+/calmodulin/Unc13A interaction plastically stabilizes vesicle priming at release sites and that CaM-domain mutation causes constitutive stabilization, thereby blocking plasticity. Labeling the functionally essential Unc13A MUN domain reveals higher STED microscopy signals closer to release sites following CaM-domain mutation. Acute phorbol ester treatment similarly enhances NT release and blocks STF/PHP in synapses expressing wild-type Unc13A, while CaM-domain mutation occludes this, indicating common downstream effects. Thus, Unc13A regulatory domains integrate signals across timescales to switch release-site participation for synaptic plasticity. [Display omitted] •Neurotransmitter release potentiation from milliseconds to minutes relies on Unc13A•Unc13A functional domains dynamically regulate release-site occupation•Phorbol esters or calmodulin binding domain mutation similarly activate Drosophila Unc13A•STED microscopy reveals nanometer changes in Unc13A at potentiated synapses Jusyte et al. find joint dependence of millisecond and minute presynaptic plasticity on conserved Unc13A. Mutating its Ca2+/calmodulin binding domain or phorbol ester treatment blocked plasticity by overactivating Unc13A, possibly by enhancing neurotransmitter release-site occupation through its subsynaptic redistribution. Converging signals on Unc13A dynamically control synaptic output from release sites.