SNAREs are concentrated in cholesterol-dependent clusters that define docking and fusion sites for exocytosis

Abstract

During exocytosis, SNARE proteins of secretory vesicles interact with the corresponding SNARE proteins in the plasmalemma to initiate the fusion reaction. However, it is unknown whether SNAREs are uniformly distributed in the membrane or whether specialized fusion sites exist. Here we report that in the plasmalemma, syntaxins are concentrated in 200 nm large, cholesterol-dependent clusters at which secretory vesicles preferentially dock and fuse. The syntaxin clusters are distinct from cholesterol-dependent membrane rafts since they are Triton X-100-soluble and do not co-patch with raft markers. Synaptosomal-associated protein (SNAP)-25 is also clustered in spots, which partially overlap with syntaxin. Cholesterol depletion causes dispersion of these clusters, which is associated with a strong reduction in the rate of secretion, whereas the characteristics of individual exocytic events are unchanged. This suggests that high local concentrations of SNAREs are required for efficient fusion.

Sdmg1 is a component of secretory granules in mouse secretory exocrine tissues

“…Lastly, the subdomains may be an in vitro artifact generated during the purification or fixation of the secretory granules. Subdomains of SNAREs and lipids within membranes have been reported in various organelles where they have been suggested to be involved in sorting membranes and cargo to different destinations, or in clustering similar proteins together to facilitate protein-protein interactions between membranes (Sonnichsen et al, 2000;Lang et al, 2001). Our data suggesting that secretory granule membranes may also contain discrete subdomains is, therefore, consistent with observations in other membrane trafficking systems.…”

Enlargeosome, an Exocytic Vesicle Resistant to Nonionic Detergents, Undergoes Endocytosis via a Nonacidic Route

“…Interestingly, this treatment had been reported to inhibit all types of regulated exocytosis investigated so far (Lang et al, 2001;Ohara-Imaizumi et al, 2004;Salaun et al, 2004). In PC12-27 cells, depletion was amply effective, as revealed by the blockade of the cholera toxin endocytosis.…”

Cholesterol transport from late endosomes to the Golgi regulates t-SNARE trafficking, assembly, and function

“…However, uneven alterations in the distribution of SNARE proteins after cholesterol depletion have been reported, ranging from moderate to complete disintegration of SNARE domains (Lang et al , 2001), indicating that lipids are important for SNARE domain integrity but that other regulatory mechanism, including additional protein–protein interactions, might also be required.…”