CAST/ELKS-endophilin-A interaction ensures synaptic vesicle pool size.

Efficient neurotransmission relies on the precise integration of synaptic vesicle (SV) recycling at the presynaptic terminal. While SV recycling is essential for neurotransmission, the molecular mechanisms coordinating vesicle dynamics at presynaptic terminals remain poorly understood. Here we report that the active zone proteins, CAST and ELKS, play a crucial role in maintaining functional SV pool size through direct interaction with endophilin-A family proteins. In cultured hippocampal neurons, disruption of CAST binding to endophilin-A resulted in a reduced number of SVs available for release, mislocalization of endophilin-A, and altered presynaptic localization of clathrin light chain. Furthermore, reduction of endophilin-A affected expression levels of active zone proteins. Collectively, these findings redefine the role of CAST/ELKS beyond active zone structural organization, demonstrating that their interaction with endophilin-A contributes to maintaining the SV pool required for sustained synaptic transmission at the presynaptic active zone.