Synaptotagmin-7 links fusion-activated Ca2+ entry (FACE) and fusion pore dilation

Abstract

Ca2+-dependent regulation of fusion pore dilation/closure is a key mechanism determining the output of cellular secretion. We have recently described „fusion-activated“ Ca2+-entry (FACE) following exocytosis of lamellar bodies (LBs) in alveolar type II cells. FACE regulates fusion pore expansion and facilitates secretion. Yet, mechanisms linking this locally restricted Ca2+ signal and fusion pore expansion were still elusive. Here we demonstrate that synaptotagmin-7 (syt-7) is expressed on LBs and links FACE and fusion pore dilation. We directly assessed dynamic changes in fusion pore diameters analysing diffusion of fluorophores across fusion pores. Expressing wt or mutant syt-7 with impaired Ca2+-binding to the C2 domains revealed that binding of Ca2+ to the C2A domain facilitates FACE-induced pore dilation, likely inhibiting translocation of complexin-2 to fused vesicles. However, the C2A domain hampered Ca2+-dependent exocytosis of LBs. These findings support that syt-7 modulates fusion pore expansion in large secretory organelles and extend our picture that LBs contain the necessary molecular inventory to facilitate secretion during the exocytic post-fusion phase. Moreover, regulating syt-7 levels on LBs appears essential to not impede exocytosis during the pre-fusion phase.