Role of the cysteine-rich domain of the t-SNARE component, SYNDET, in membrane binding and subcellular localization

Wild-type syndet is efficiently recruited at the plasma membrane in transfected AtT-20 cells. A deletion at the cysteine-rich domain abolishes palmitoylation, membrane binding, and plasma membrane distribution of syndet. Syndet, SNAP-25A, and SNAP-25B share four cysteine residues, of which three, Cys(2), Cys(4), and Cys(5), are absolutely conserved in all three homologs, Mutations at any pair of cysteines within cysteines 2, 4, and 5 shift syndet from the cell surface into the cytoplasm, Thus, at least two cysteines within the conserved triplet are necessary for plasma membrane localization. Syndet C1S/C3S, with substitutions at the pair Cys(1) and Cys(3), distributes to the plasma membrane, a Gels-like compartment, and the cytosol, We conclude that Cys(1) and Cys(3) are not absolutely necessary for membrane binding or plasma membrane localization. Our results show that the cysteine-rich domain of syndet plays a major role in its subcellular distribution.