t-SNARE cluster organization and dynamics in chromaffin cells

Adrenomedullary chromaffin cells represent an excellent model to study the molecular events linked to exocytosis, because they use the same type of SNAREs for vesicle docking and fusion as neurons. In these cells, both in the intact tissue and in isolated cells in culture, syntaxin-1 and SNAP-25 are present in the plasmalemma unevenly distributed in patches, even when exogenous t-SNAREs are expressed. In fact, the expression of SNAP-25 fused to green fluorescent protein has been useful to study the movement of these clusters by total internal reflection fluorescent microscopy. These microdomains move little in the plasma membrane plane but they undertake relatively large displacements of 100 nm in the axis perpendicular to the membrane. Movement in either axis is dependent on molecular interactions within the t-SNARE complex and indeed, clusters formed by recombinant SNAP-25 Δ9, the product of Botulinum neurotoxin A cleavage, undergo larger displacement. Interestingly, altering the movement of t-SNARE clusters also influences the mobility of the chromaffin vesicles associated with these t-SNAREs. Furthermore, highly mobile vesicles associated with the clusters formed by SNAP-25 Δ9 present a low probability of exocytosis and also slower fusion kinetics. Finally, we discuss some of the factors that could influence the movement of t-SNARE clusters and how these dynamics may influence the mobility and the fusion properties of the vesicles in the vicinity of active sites.