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

During exocytosis, SNARE vesicles interact with the corresponding SNARE pro teins in the plasmalemma to initiate the fusion reaction. However, it is un known whether SNAREs are uniformly distributed in the membrane or whether s pecialized fusion sites exist. Here we report that in the plasmalemma, synt axins are concentrated in 200 nm large, cholesterol-dependent clusters at w hich secretory vesicles preferentially dock and fuse. The syntaxin clusters are distinct from cholesterol-dependent membrane rafts since they are Trit on X-100-soluble and do not co-patch with raft markers. Synaptosomal-associ ated protein (SNAP)-25 is also clustered in spots, which partially overlap with syntaxin. Cholesterol depletion causes dispersion of these clusters, w hich is associated with a strong reduction in the rate of secretion, wherea s the characteristics of individual exocytic events are unchanged. This sug gests that high local concentrations of SNAREs are required for efficient f usion.