Hemi-fused structure mediates and controls fusion and fission in live cells

Abstract

Membrane fusion and fission are vital to eukaryotes’ life1–5. For three decades, it has been proposed that fusion is mediated by fusion between proximal leaflets of two bilayers (hemi-fusion) that produces a hemi-fused structure, followed by fusion between distal leaflets, whereas fission is via hemi-fission, which also produces a hemi-fused structure, followed by full fission1, 4, 6–10. This hypothesis remained unsupported owing to the lack of observation of hemi-fusion/hemi-fission in live cells. A competing fusion hypothesis involving protein-lined pore formation has also been proposed2, 11–15. Using confocal and super-resolution STED microscopy, we observed the hemi-fused Ω-shaped structure for the first time in live cells, neuroendocrine chromaffin cells and pancreatic β-cells. This structure was generated from fusion pore opening or closure (fission) at the plasma membrane. Unexpectedly, its transition to full fusion or fission was determined by competition between fusion and calcium/dynamin-dependent fission mechanisms, and was surprisingly slow (seconds to tens of seconds) in a significant fraction of the events. These results provide key missing evidence over the past three decades proving the hemi-fusion and hemi-fission hypothesis in live cells, and reveal the hemi-fused intermediate as a key structure controlling fusion/fission, as fusion and fission mechanisms compete to determine its transition to fusion or fission.

High-resolution imaging of cellular dopamine efflux using a fluorescent nanosensor array

“…S17 and S18). Total internal reflection fluorescence microscopy studies of labeled vesicles demonstrated exocytosis from the bottom of chromaffine cells (45). However, these methods naturally focus on the parts of the cell close to the substrate (i.e., cell bottom).…”

The Rab3A-22A Chimera Prevents Sperm Exocytosis by Stabilizing Open Fusion Pores

“…Fusion generates an ⍀-shaped membrane profile. It was believed that the pore either dilated (full collapse) or resealed (kiss and run), but the existence of a larger spectrum of membrane fates has been recently unveiled (6,7).…”

Real-time insights into regulated exocytosis

“…This mode of secretion of large mucinous cargo is considerably different from that of hormone secretion in endocrine cells as demonstrated recently (Zhao et al, 2016). Confocal and super-resolution stimulated emission depletion (STED) imaging of secretion in chromaffin cells and pancreatic β-cells has revealed a hemi-fusion intermediate between the secretory vesicle and the PM (Zhao et al, 2016), where fusion between the inner leaflet of the cell and the outer lipid layer of the secretory granule forms in the absence of a complete fusion pore. Interestingly, these intermediates were stable for many seconds before full fusion occurred, suggesting that secretion proceeds through a hemi-fused intermediate.…”