Targeting of SNAP-23 and SNAP-25 in Polarized Epithelial Cells*
- Seng Hui Low‡§,
- Paul A. Roche¶,
- Howard A. Anderson¶,
- Sven C. D. van Ijzendoorn‡‖**,
- Min Zhang‡,
- Keith E. Mostov‡ and
- Thomas Weimbs‡§‡
- From the ‡Department of Anatomy, Department of Biochemistry and Biophysics, Cardiovascular Research Institute, University of California, San Francisco, California 94143-0452,¶Experimental Immunology Branch, NCI, National Institutes of Health, Bethesda, Maryland 20892, ‖Department of Physiological Chemistry, Groningen University, 9713 AV Groningen, The Netherlands
Abstract
SNAP-23 is the ubiquitously expressed homologue of the neuronal SNAP-25, which functions in synaptic vesicle fusion. We have investigated the subcellular localization of SNAP-23 in polarized epithelial cells. In hepatocyte-derived HepG2 cells and in Madin-Darby canine kidney (MDCK) cells, the majority of SNAP-23 was present at both the basolateral and apical plasma membrane domains with little intracellular localization. This suggests that SNAP-23 does not function in intracellular fusion events but rather as a general plasma membrane t-SNARE. Canine SNAP-23 is efficiently cleaved by the botulinum neurotoxin E, suggesting that it is the toxin-sensitive factor previously found to be involved in plasma membrane fusion in MDCK cells. The localization of SNAP-25 in transfected MDCK cells was studied for comparison and was found to be identical to SNAP-23 with the exception that SNAP-25 was transported to the primary cilia protruding from the apical plasma membrane, which suggests that subtle differences in the targeting signals of both proteins exist. In contrast to its behavior in neurons, the distribution of SNAP-25 in MDCK cells remained unaltered by treatment with dibutyryl cAMP or forskolin, which, however, caused an increased growth of the primary cilia. Finally, we found that SNAP-23/25 and syntaxin 1A, when co-expressed in MDCK cells, do not stably interact with each other but are independently targeted to the plasma membrane and lysosomes, respectively.
Footnotes
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↵* This work was supported by National Institutes of Health Grants R01 AI25144, AI39161, and AI36953 (to K. E. M.).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 Irvington Institute for Immunology.
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↵** Supported by a travel grant from the Groningen Institute for Drug Studies.
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↵‡ Supported by a Feodor-Lynen-Fellowship of the Alexander von Humboldt-Foundation. To whom correspondence should be addressed: Dept. of Anatomy, University of California San Francisco, 513 Parnassus Ave., San Francisco, CA 94143-0452. E-mail: weimbs{at}itsa.ucsf.edu.
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↵1 The abbreviations used are: SNAP-25, synaptosomal-associated protein of 25 kDa; NSF,N-ethylmaleimide-sensitive fusion protein; BC, bile canaliculi; Bt2cAMP, dibutyryl cAMP; MDCK, Madin-Darby canine kidney; FBS, fetal bovine serum; PAGE, polyacrylamide gel electrophoresis.
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- Received July 3, 1997.
- Revision received November 12, 1997.
- The American Society for Biochemistry and Molecular Biology, Inc.
