The synaptotagmin C2B domain calcium-binding loops modulate the rate of fusion pore expansion

Journal Article · Sun Apr 01 00:00:00 EDT 2018 · Molecular Biology of the Cell

DOI:https://doi.org/10.1091/mbc.E17-11-0623· OSTI ID:1544301

Bendahmane, Mounir ^[1]; Bohannon, Kevin P. ^[1]; Bradberry, Mazdak M. ^[2]; Rao, Tejeshwar C. ^[1]; Schmidtke, Michael W. ^[1]; Abbineni, Prabhodh S. ^[1]; Chon, Nara L. ^[3]; Tran, Sherleen ^[3]; Lin, Hai ^[3]; Chapman, Edwin R. ^[2]; Knight, Jefferson D. ^[3]; Anantharam, Arun ^[1]; Brennwald, Patrick J.

---

1. Department of Pharmacology, University of Michigan, Ann Arbor, MI 48109
2. Department of Neuroscience, Howard Hughes Medical Institute. University of Wisconsin–Madison, Madison, WI 53705
3. Department of Chemistry, University of Colorado Denver, Denver, CO 80217

In chromaffin cells, the kinetics of fusion pore expansion vary depending on which synaptotagmin isoform (Syt-1 or Syt-7) drives release. Our recent studies have shown that fusion pores of granules harboring Syt-1 expand more rapidly than those harboring Syt-7. Here we sought to define the structural specificity of synaptotagmin action at the fusion pore by manipulating the Ca²⁺-binding C2B module. We generated a chimeric Syt-1 in which its C2B Ca²⁺-binding loops had been exchanged for those of Syt-7. Fusion pores of granules harboring a Syt-1 C2B chimera with all three Ca²⁺-binding loops of Syt-7 (Syt-1:7C2B₁₂₃) exhibited slower rates of fusion pore expansion and neuropeptide cargo release relative to WT Syt-1. After fusion, this chimera also dispersed more slowly from fusion sites than WT protein. We speculate that the Syt-1:7 C2B₁₂₃and WT Syt-1 are likely to differ in their interactions with Ca²⁺and membranes. Subsequent in vitro and in silico data demonstrated that the chimera exhibits a higher affinity for phospholipids than WT Syt-1. We conclude that the affinity of synaptotagmin for the plasma membrane, and the rate at which it releases the membrane, contribute in important ways to the rate of fusion pore expansion.

• Research Organization: Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). National Energy Research Scientific Computing Center (NERSC)
• Sponsoring Organization: USDOE
• OSTI ID: 1544301
• Journal Information: Molecular Biology of the Cell, Vol. 29, Issue 7; ISSN 1059-1524
• Publisher: American Society for Cell Biology
• Country of Publication: United States
• Language: English