The tandem C2 domains of synaptotagmin contain redundant Ca2+ binding sites that cooperate to engage t-SNAREs and trigger exocytosis

doi:10.1083/jcb.200105020

Published online 10 September 2001. doi:10.1083/jcb.200105020

This Article

	Full Text
	PDF (Full Text)
	Alert me when this article is cited
	Citation Map

Services

	Email this article
	Similar articles in this journal
	Similar articles in PubMed
	Alert me to new content in the JCB
	Download to citation manager

Citing Articles

	Citing Articles via HighWire
	Citing Articles via CrossRef
	Citing Articles via Google Scholar

Google Scholar

	Articles by Earles, C. A.
	Articles by Chapman, E. R.
	Search for Related Content

PubMed

	PubMed Citation
	Articles by Earles, C. A.
	Articles by Chapman, E. R.


Social Bookmarking

	What's this?

© The Rockefeller University Press, 0021-9525/2001/9/1117 $5.00
The Journal of Cell Biology, Volume 154, Number 6, September 17, 2001 1117-1124

Report

The tandem C2 domains of synaptotagmin contain redundant Ca²+ binding sites that cooperate to engage t-SNAREs and trigger exocytosis

Cynthia A. Earles, Jihong Bai, Ping Wang and Edwin R. Chapman

Department of Physiology, University of Wisconsin, Madison, WI 53706

Address correspondence to Edwin R. Chapman, Dept. of Physiology, SMI 129, University of Wisconsin, 1300 University Ave., Madison, WI 53706. Tel.: (608) 263-1762. Fax: (608) 265-5512. E-mail: chapman{at}physiology.wisc.edu

Real-time voltammetry measurements from cracked PC12 cells wereused to analyze the role of synaptotagmin–SNARE interactionsduring Ca²+-triggered exocytosis. The isolated C2A domain ofsynaptotagmin I neither binds SNAREs nor inhibits norepinephrinesecretion. In contrast, two C2 domains in tandem (either C2A-C2Bor C2A-C2A) bind strongly to SNAREs, displace native synaptotagminfrom SNARE complexes, and rapidly inhibit exocytosis. The tandemC2 domains of synaptotagmin cooperate via a novel mechanismin which the disruptive effects of Ca²⁺ ligand mutations inone C2 domain can be partially alleviated by the presence ofan adjacent C2 domain. Complete disruption of Ca²⁺-triggeredmembrane and target membrane SNARE interactions required simultaneousneutralization of Ca²⁺ ligands in both C2 domains of the protein.We conclude that synaptotagmin–SNARE interactions regulatemembrane fusion and that cooperation between synaptotagmin'sC2 domains is crucial to its function.

Key Words: synaptotagmin; SNARE; membrane fusion; C2 domain; exocytosis

Add to CiteULike CiteULike Add to Complore Complore Add to Connotea Connotea Add to Del.icio.us Del.icio.us Add to Digg Digg Add to Reddit Reddit Add to Technorati Technorati What's this?

This article has been cited by other articles:

Zhang, Z., Jackson, M. B. (2008). Temperature Dependence of Fusion Kinetics and Fusion Pores in Ca2+-triggered Exocytosis from PC12 Cells. J. Gen. Physiol. 131: 117-124 [Abstract] [Full Text]
Lynch, K. L., Gerona, R.R.L., Larsen, E. C., Marcia, R. F., Mitchell, J. C., Martin, T.F.J. (2007). Synaptotagmin C2A Loop 2 Mediates Ca2+-dependent SNARE Interactions Essential for Ca2+-triggered Vesicle Exocytosis. Mol. Biol. Cell 18: 4957-4968 [Abstract] [Full Text]
Gattegno, T., Mittal, A., Valansi, C., Nguyen, K. C.Q., Hall, D. H., Chernomordik, L. V., Podbilewicz, B. (2007). Genetic Control of Fusion Pore Expansion in the Epidermis of Caenorhabditis elegans. Mol. Biol. Cell 18: 1153-1166 [Abstract] [Full Text]
Czibener, C., Sherer, N. M., Becker, S. M., Pypaert, M., Hui, E., Chapman, E. R., Mothes, W., Andrews, N. W. (2006). Ca2+ and synaptotagmin VII-dependent delivery of lysosomal membrane to nascent phagosomes. J. Cell Biol. 174: 997-1007 [Abstract] [Full Text]
Aganna, E., Burrin, J. M, Hitman, G. A, Turner, M. D (2006). Involvement of calpain and synaptotagmin Ca2+ sensors in hormone secretion from excitable endocrine cells.. J Endocrinol 190: R1-R7 [Abstract] [Full Text]
Hui, E., Bai, J., Chapman, E. R. (2006). Ca2+-Triggered Simultaneous Membrane Penetration of the Tandem C2-Domains of Synaptotagmin I. Biophys. J 91: 1767-1777 [Abstract] [Full Text]
Ahras, M., Otto, G. P., Tooze, S. A. (2006). Synaptotagmin IV is necessary for the maturation of secretory granules in PC12 cells. J. Cell Biol. 173: 241-251 [Abstract] [Full Text]
Wang, C.-T., Bai, J., Chang, P. Y., Chapman, E. R., Jackson, M. B. (2006). Synaptotagmin-Ca2+ triggers two sequential steps in regulated exocytosis in rat PC12 cells: fusion pore opening and fusion pore dilation. J. Physiol. 570: 295-307 [Abstract] [Full Text]
Rhee, J.-S., Li, L. Y., Shin, O.-H., Rah, J.-C., Rizo, J., Sudhof, T. C., Rosenmund, C. (2005). Augmenting neurotransmitter release by enhancing the apparent Ca2+ affinity of synaptotagmin 1. Proc. Natl. Acad. Sci. USA 102: 18664-18669 [Abstract] [Full Text]
Wang, P., Chicka, M. C., Bhalla, A., Richards, D. A., Chapman, E. R. (2005). Synaptotagmin VII Is Targeted to Secretory Organelles in PC12 Cells, Where It Functions as a High-Affinity Calcium Sensor. Mol. Cell. Biol. 25: 8693-8702 [Abstract] [Full Text]
Bhalla, A., Tucker, W. C., Chapman, E. R. (2005). Synaptotagmin Isoforms Couple Distinct Ranges of Ca2+, Ba2+, and Sr2+ Concentration to SNARE-mediated Membrane Fusion. Mol. Biol. Cell 16: 4755-4764 [Abstract] [Full Text]
DeBlasio, S. L., Luesse, D. L., Hangarter, R. P. (2005). A Plant-Specific Protein Essential for Blue-Light-Induced Chloroplast Movements. Plant Physiol. 139: 101-114 [Abstract] [Full Text]
Shen, S. S., Tucker, W. C., Chapman, E. R., Steinhardt, R. A. (2005). Molecular Regulation of Membrane Resealing in 3T3 Fibroblasts. J. Biol. Chem. 280: 1652-1660 [Abstract] [Full Text]
Maher, B. J., MacKinnon, R. L. II, Bai, J., Chapman, E. R., Kelly, P. T. (2005). Activation of Postsynaptic Ca2+ Stores Modulates Glutamate Receptor Cycling in Hippocampal Neurons. J. Neurophysiol. 93: 178-188 [Abstract] [Full Text]
Llinas, R. R., Sugimori, M., Moran, K. A., Moreira, J. E., Fukuda, M. (2004). Vesicular reuptake inhibition by a synaptotagmin I C2B domain antibody at the squid giant synapse. Proc. Natl. Acad. Sci. USA 101: 17855-17860 [Abstract] [Full Text]
Yoshihara, M., Montana, E. S. (2004). The Synaptotagmins: Calcium Sensors for Vesicular Trafficking. Neuroscientist 10: 566-574 [Abstract]
Tucker, W. C., Weber, T., Chapman, E. R. (2004). Reconstitution of Ca2+-Regulated Membrane Fusion by Synaptotagmin and SNAREs. Science 304: 435-438 [Abstract] [Full Text]
Chieregatti, E., Chicka, M. C., Chapman, E. R., Baldini, G. (2004). SNAP-23 Functions in Docking/Fusion of Granules at Low Ca2+. Mol. Biol. Cell 15: 1918-1930 [Abstract] [Full Text]
Wang, P., Wang, C.-T., Bai, J., Jackson, M. B., Chapman, E. R. (2003). Mutations in the Effector Binding Loops in the C2A and C2B Domains of Synaptotagmin I Disrupt Exocytosis in a Nonadditive Manner. J. Biol. Chem. 278: 47030-47037 [Abstract] [Full Text]
Edwardson, J. M., Wang, C.-T., Gong, B., Wyttenbach, A., Bai, J., Jackson, M. B., Chapman, E. R., Morton, A. J. (2003). Expression of Mutant Huntingtin Blocks Exocytosis in PC12 Cells by Depletion of Complexin II. J. Biol. Chem. 278: 30849-30853 [Abstract] [Full Text]
Duman, J. G., Forte, J. G. (2003). What is the role of SNARE proteins in membrane fusion?. Am. J. Physiol. Cell Physiol. 285: C237-C249 [Abstract] [Full Text]
Tucker, W. C., Edwardson, J. M., Bai, J., Kim, H.-J., Martin, T. F.J., Chapman, E. R. (2003). Identification of synaptotagmin effectors via acute inhibition of secretion from cracked PC12 cells. J. Cell Biol. 162: 199-209 [Abstract] [Full Text]
Wu, Y., He, Y., Bai, J., Ji, S.-R., Tucker, W. C., Chapman, E. R., Sui, S.-F. (2003). Visualization of synaptotagmin I oligomers assembled onto lipid monolayers. Proc. Natl. Acad. Sci. USA 100: 2082-2087 [Abstract] [Full Text]
Fukuda, M., Kanno, E., Ogata, Y., Saegusa, C., Kim, T., Loh, Y. P., Yamamoto, A. (2003). Nerve Growth Factor-dependent Sorting of Synaptotagmin IV Protein to Mature Dense-core Vesicles That Undergo Calcium-dependent Exocytosis in PC12 Cells. J. Biol. Chem. 278: 3220-3226 [Abstract] [Full Text]
Fernandez-Chacon, R., Shin, O.-H., Konigstorfer, A., Matos, M. F., Meyer, A. C., Garcia, J., Gerber, S. H., Rizo, J., Sudhof, T. C., Rosenmund, C. (2002). Structure/Function Analysis of Ca2+ Binding to the C2A Domain of Synaptotagmin 1. J. Neurosci. 22: 8438-8446 [Abstract] [Full Text]
Fukuda, M. (2002). Vesicle-associated Membrane Protein-2/Synaptobrevin Binding to Synaptotagmin I Promotes O-Glycosylation of Synaptotagmin I. J. Biol. Chem. 277: 30351-30358 [Abstract] [Full Text]
Mahal, L. K., Sequeira, S. M., Gureasko, J. M., Sollner, T. H. (2002). Calcium-independent stimulation of membrane fusion and SNAREpin formation by synaptotagmin I. J. Cell Biol. 158: 273-282 [Abstract] [Full Text]
Saegusa, C., Fukuda, M., Mikoshiba, K. (2002). Synaptotagmin V Is Targeted to Dense-core Vesicles That Undergo Calcium-dependent Exocytosis in PC12 Cells. J. Biol. Chem. 277: 24499-24505 [Abstract] [Full Text]
Honda, A., Yamada, M., Saisu, H., Takahashi, H., Mori, K. J., Abe, T. (2002). Direct, Ca2+-dependent Interaction between Tubulin and Synaptotagmin I. A POSSIBLE MECHANISM FOR ATTACHING SYNAPTIC VESICLES TO MICROTUBULES. J. Biol. Chem. 277: 20234-20242 [Abstract] [Full Text]
Bai, J., Wang, P., Chapman, E. R. (2002). C2A activates a cryptic Ca2+-triggered membrane penetration activity within the C2B domain of synaptotagmin I. Proc. Natl. Acad. Sci. USA 10.1073/pnas.032541099v1 [Abstract] [Full Text]
Bai, J., Wang, P., Chapman, E. R. (2002). C2A activates a cryptic Ca2+-triggered membrane penetration activity within the C2B domain of synaptotagmin I. Proc. Natl. Acad. Sci. USA 99: 1665-1670 [Abstract] [Full Text]