Synaptojanin Forms Two Separate Complexes in the Nerve Terminal
INTERACTIONS WITH ENDOPHILIN AND AMPHIPHYSIN*
- From the ‡Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, 3801 Rue University, Montreal, Quebec H3A 2B4, Canada and the¶Department of Biology, University of North Carolina, Chapel Hill, North Carolina 27599
Abstract
Endophilin is a recently discovered src homology 3 domain-containing protein that is a major in vitrobinding partner for synaptojanin. To further characterize endophilin, we generated an antipeptide antibody. Endophilin is enriched in the brain, and immunofluorescence analysis reveals a high concentration of the protein in synaptic terminals, where it colocalizes with synaptojanin. In vitro binding assays demonstrate that endophilin binds through its src homology 3 domain to synaptojanin, and immunoprecipitation analysis with the antiendophilin antibody reveals that endophilin is stably associated with synaptojanin in the nerve terminal. Immunoprecipitation with an antibody against amphiphysin I and II, which interact through their src homology 3 domains with dynamin and synaptojanin at sites distinct from those for endophilin, reveals a second stable complex, which includes dynamin and synaptojanin but excludes endophilin. These data demonstrate that synaptojanin is present in two separate complexes in the nerve terminal and support an important role for endophilin in the regulation of synaptojanin function.
Footnotes
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↵* This work was supported by Grants MT-13461 from the Medical Research Council of Canada and 197685 from the Natural Sciences and Engineering Research Council (to P. S. 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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↵§ Medical Research Council of Canada Postdoctoral Fellow.
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↵‖ Scholar of the Medical Research Council of Canada and an Alfred P. Sloan Research Fellow. To whom correspondence should be addressed. Tel.: 514-398-7355; Fax: 514-398-8106; E-mail:mcpm{at}musica.mcgill.ca.
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↵1 The abbreviations used are: PCR, polymerase chain reaction; GST, glutathione S-transferase; PAGE, polyacrylamide gel electrophoresis.
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- Received June 10, 1997.
- Revision received August 5, 1997.
