The synaptic vesicle protein, cysteine-string protein, is associated with the plasma membrane in 3T3-L1 adipocytes and interacts with syntaxin 4

Adipocytes and muscle cells play a major role in blood glucose homeostasis. This is dependent upon the expression of Glut4, an insulin-responsive faci litative glucose transporter. Glut4 is localised to specialised intracellul ar vesicles that fuse with the plasma membrane in response to insulin stimu lation, The insulin-induced translocation of Glut4 to the cell surface is e ssential for the maintenance of optimal blood glucose levels, and defects i n this system are associated with insulin resistance and type II diabetes. Therefore, a major focus of recent research has been to identify and charac terise proteins that regulate Glut4 translocation. Cysteine-string protein (Csp) is a secretory vesicle protein that functions in presynaptic neurotra nsmission and also in regulated exocytosis from non-neuronal cells. We show that Csp1 is expressed in 3T3-L1 adipocytes and that cellular levels of th is protein are increased following cell differentiation. Combined fractiona tion and immunofluorescence analyses reveal that Csp1 is not a component of intracellular Glut4-storage vesicles (GSVs), but is associated with the ad ipocyte plasma membrane. This association is stable, and not affected by ei ther insulin stimulation or chemical depalmitoylation of Csp1. We also demo nstrate that Csp1 interacts with the t-SNARE syntaxin 4, As syntaxin 4 is a n important mediator of insulin-stimulated GSV fusion with the plasma membr ane, this suggests that Csp1 may play a regulatory role in this process. Sy ntaxin 4 interacts specifically with Csp1, but not with Csp2, In contrast, syntaxin 1A binds to both Csp isoforms, and actually exhibits a higher affi nity for the Csp2 protein. The results described raise a number of interesting questions concerning th e intracellular targeting of Csp in different cell types, and suggest that the composition and synthesis of GSVs may be different from synaptic and ot her secretory vesicles, In addition, the interaction of Csp1 with syntaxin 4 suggests that this Csp isoform may play a role in insulin-stimulated fusi on of GSVs with the plasma membrane.