Need for speed: Super-resolving the dynamic nanoclustering of syntaxin-1 at exocytic fusion sites

Abstract

Communication between cells relies on regulated exocytosis, a multi-step process that involves the docking, priming and fusion of vesicles with the plasma membrane, culminating in the release of neurotransmitters and hormones. Key proteins and lipids involved in exocytosis are subjected to Brownian movement and constantly switch between distinct motion states which are governed by short-lived molecular interactions. Critical biochemical reactions between exocytic proteins that occur in the confinement of nanodomains underpin the precise sequence of priming steps which leads to the fusion of vesicles. The advent of super-resolution microscopy techniques has provided the means to visualize individual molecules on the plasma membrane with high spatiotemporal resolution in live cells. These techniques are revealing a highly dynamic nature of the nanoscale organization of the exocytic machinery. In this review, we focus on soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) syntaxin-1, which mediates vesicular fusion. Syntaxin-1 is highly mobile at the plasma membrane, and its inherent speed allows fast assembly and disassembly of syntaxin-1 nanoclusters which are associated with exocytosis. We reflect on recent studies which have revealed the mechanisms regulating syntaxin-1 nanoclustering on the plasma membrane and draw inferences on the effect of synaptic activity, phosphoinositides, N-ethylmaleimide-sensitive factor (NSF), -soluble NSF attachment protein (-SNAP) and SNARE complex assembly on the dynamic nanoscale organization of syntaxin-1.Abbreviations: SNARE, soluble N-ethylmaleimide-sensitive factor attachment receptor; NSF, N-ethylmaleimide-sensitive factor; -SNAP, -soluble NSF attachment protein; PC12, pheochromocytoma; NMJ, neuromuscular junction; TIRF, total internal reflection fluorescence; STED, stimulated emission depletion; dSTORM, direct stochastic optical resolution microscopy; PALM, photoactivated localization microscopy; SPT, single particle tracking; sptPALM, single particle tracking photoactivated localization microscopy; uPAINT, universal point accumulation in nanoscale topography; PtdIns(4,5)P 2 , phosphatidylinositol (4,5)-biphosphate; PtdIns(3,4,5)P 3 , phosphatidylinositol (3,4,5)-biphosphate; FRAP, fluorescence recovery after bleaching; GFP, green fluorescent protein. Highlights-Syntaxin-1 forms nanoclusters on the plasma membrane.-Multiple molecular mechanisms regulate syntaxin-1 nanoclustering on the plasma membrane.-Synaptic activity, phosphoinositides and SNARE complex assembly differentially control the nanoscale organization of syntaxin-1 on the plasma membrane.-Super-resolution microscopy has the potential to uncover the design principles governing regulated exocytosis.

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