Interactive nanocluster compaction of the ELKS scaffold and Cacophony Ca2+channels drives sustained active zone potentiation

At presynaptic active zones (AZs), conserved scaffold protein architectures control synaptic vesicle (SV) release by defining the nanoscale distribution and density of voltage-gated Ca2+ channels (VGCCs). While AZs can po-tentiate SV release in the minutes range, we lack an understanding of how AZ scaffold components and VGCCs engage into potentiation. We here establish dynamic, intravital single-molecule imaging of endogenously tagged proteins at Drosophila AZs undergoing presynaptic homeostatic potentiation. During potentiation, the numbers of alpha 1 VGCC subunit Cacophony (Cac) increased per AZ, while their mobility decreased and nano -scale distribution compacted. These dynamic Cac changes depended on the interaction between Cac channel's intracellular carboxyl terminus and the membrane-close amino-terminal region of the ELKS-family protein Bruchpilot, whose distribution compacted drastically. The Cac-ELKS/Bruchpilot interaction was also needed for sustained AZ potentiation. Our single-molecule analysis illustrates how the AZ scaffold couples to VGCC nanoscale distribution and dynamics to establish a state of sustained potentiation.

Automated summary

In this study, dynamic intravital single-molecule imaging was used to investigate the interaction between scaffold proteins and voltage-gated calcium channels (VGCCs) during presynaptic homeostatic potentiation in Drosophila AZs. It was found that the interaction between the intracellular carboxyl terminus of the Cac channel and the amino-terminal region of the ELKS-family protein Bruchpilot played a crucial role in the sustained potentiation of AZs.