Role of the Cysteine-rich Domain of the t-SNARE Component, SYNDET, in Membrane Binding and Subcellular Localization*
- From the Department of Anatomy and Cell Biology, Columbia University, College of Physicians and Surgeons, New York, New York 10032
Abstract
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, Cys2, Cys4, and Cys5, 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 Cys1 and Cys3, distributes to the plasma membrane, a Golgi-like compartment, and the cytosol. We conclude that Cys1 and Cys3 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.
Footnotes
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↵* This work was supported in part by National Institutes of Health Grant RO1-DK53293 and a grant-in-aid from the American Heart Association, New York City Affiliate (to G. B. S.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ Supported by the Naomie Berrie Diabetes Fellowship at Columbia Presbyterian Medical Center.
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↵§ To whom correspondence should be addressed. Tel.: 212-305-6405; Fax: 212-305-3970; E-mail: gb74{at}columbia.edu.
- Abbreviations:
- PCR
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polymerase chain reaction
- PAGE
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polyacrylamide gel electrophoresis
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- Received July 20, 1998.
- Revision received December 21, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
