Distinct Protein Domains Are Responsible for the Interaction of Hrs-2 with SNAP-25
THE ROLE OF Hrs-2 IN 7 S COMPLEX FORMATION*
- From the Department of Neurobiology and Anatomy and the W. M. Keck Center for the Neurobiology of Learning and Memory, University of Texas Medical School, Houston, Texas 77030
Abstract
Regulated secretion of neurotransmitter at the synapse is likely to be mediated by dynamic protein interactions involving components of the vesicle (vesicle-associated membrane protein; VAMP) and plasma membrane (syntaxin and synaptosomal associated protein of 25 kDa (SNAP-25)) along with additional molecules that allow for the regulation of this process. Recombinant Hrs-2 interacts with SNAP-25 in a calcium-dependent manner (they dissociate at elevated calcium levels) and inhibits neurotransmitter release. Thus, Hrs-2 has been hypothesized to serve a negative regulatory role in secretion through its interaction with SNAP-25. In this report, we show that Hrs-2 and SNAP-25 interact directly through specific coiled-coil domains in each protein. The presence of syntaxin enhances the binding of Hrs-2 to SNAP-25. Moreover, while both Hrs-2 and VAMP can separately bind to SNAP-25, they cannot bind simultaneously. Additionally, the presence of Hrs-2 reduces the incorporation of VAMP into the syntaxin·SNAP-25·VAMP (7 S) complex. These findings suggest that Hrs-2 may modulate exocytosis by regulating the assembly of a protein complex implicated in membrane fusion.
Footnotes
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↵* This work was supported in part by the Mallinckrodt Foundation and National Institutes of Health Grant MH058920.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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↵‡ To whom correspondence should be addressed: Dept. of Neurobiology and Anatomy, University of Texas Medical School, 6431 Fannin St., Rm. 7.208, Houston, TX 77030. Tel.: 713-500-5614; Fax: 713-500-0623; E-mail: abean@nba19.med.uth.tmc.edu.
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↵2 S. Tsujimoto and A.J. Bean, unpublished observations.
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↵3 S. Davanger and A.J. Bean, unpublished observations.
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↵4 A. J. Bean, unpublished observations.
- Abbreviations:
- SNAP-25
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synaptosomal associated protein of 25 kDa
- VAMP
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vesicle-associated membrane protein
- Hrs-2
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hepatocyte growth factor-regulated tyrosine kinase substrate 2
- GST
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glutathione S-transferase
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- Received August 2, 1999.
- Revision received November 4, 1999.
- The American Society for Biochemistry and Molecular Biology, Inc.
