SNAP-25 Traffics to the Plasma Membrane by a Syntaxin-independent Mechanism

• Published in: The Journal of biological chemistry 2002-09, Vol.277 (37), p.34303-34309
• Main Authors: Loranger, Stephanie S., Linder, Maurine E.
• Format: Article
• Language: English
• Subjects: Biological Transport; Cell Membrane - metabolism; Cytoplasm - metabolism; HeLa Cells; Humans; Membrane Proteins - chemistry; Membrane Proteins - metabolism; Membrane Proteins - physiology; Nerve Tissue Proteins - chemistry; Nerve Tissue Proteins - metabolism; Palmitic Acid - metabolism; Protein Structure, Secondary; Qa-SNARE Proteins; Synaptosomal-Associated Protein 25; Syntaxin 1
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M202125200

SNAREs (solubleN-ethylmaleimide-sensitive factorattachment protein receptors) are essential for vesicle docking and fusion. SNAP-25, syntaxin 1A, and synaptobrevin/vesicle-associated membrane protein (VAMP) are SNARE proteins that mediate fusion of synaptic vesicles with the plasma membrane. It has been proposed that interactions of SNAP-25 with syntaxin 1A are required for initial membrane attachment of SNAP-25 (Vogel, K., Cabaniols, J.-P., and Roche, P. (2000)J. Biol. Chem. 275, 2959–2965). However, we have shown previously that residues 85–120 of the SNAP-25 interhelical domain, which do not interact with syntaxin, are necessary and sufficient for palmitoylation and plasma membrane localization of a green fluorescent protein reporter molecule (Gonzalo, S., Greentree, W. K., and Linder, M. E. (1999) J. Biol. Chem. 274, 21313–21318). To clarify the role of syntaxin in membrane targeting of SNAP-25, we studied a SNAP-25 point mutant (G43D) that does not interact with syntaxin. SNAP-25 G43D/green fluorescent protein was palmitoylated and localized at the plasma membrane. Newly synthesized SNAP-25 G43D had the same kinetics of membrane association as the wild-type protein. Furthermore, expression of a cytosolic mutant syntaxin 1A did not interfere with SNAP-25 membrane interactions or palmitoylation in the neuronal cell line NG108-15. Exogenously expressed SNAP-25 targets efficiently to the plasma membrane in cells of neuronal origin but only partially in HeLa cells, a neurosecretion-incompetent line. This phenotype was not rescued when syntaxin 1A was co-expressed with SNAP-25. Our data support a syntaxin-independent mechanism of membrane targeting for SNAP-25.