Fig. 10. Schematic model of GAD65 cycling between the cytosol, the ER-Golgi and a vesicular pathway. Newly synthesized hydrophilic and soluble GAD65 (F1) undergoes an irreversible hydrophobic modification in the cytosol (Step A). The resulting hydrophobic GAD65 (F2) reversibly associates with ER and Golgi membranes, establishing an equilibrium between membrane and cytosolic pools. In Golgi membranes, the protein undergoes palmitoylation (Step B) catalyzed by a palmitoyl transferase (PAT). The palmitoylated protein (F3) is targeted to the TGN, shifting the equilibrium from the ER towards the Golgi. From the TGN, the protein is targeted to cytosolic vesicles and a pathway to synaptic vesicles and presynaptic termini in neurons and synaptic-like microvesicles in pancreatic β cells. A depalmitoylation along this pathway (Step C) results in a reversal to F2, trafficking back to ER-Golgi membranes by a microtubule-independent pathway, and a new cycle of palmitoylation and targeting to the vesicular pathway.