SYNAPTOJANIN FORMS 2 SEPARATE COMPLEXES IN THE NERVE-TERMINAL - INTERACTIONS WITH ENDOPHILIN AND AMPHIPHYSIN

Endophilin is a recently discovered src homology 3 domain-containing p rotein that is a major in vitro binding partner for synaptojanin. To f urther characterize endophilin, we generated an antipeptide antibody. Endophilin is enriched in the brain, and immunofluorescence analysis r eveals a high concentration of the protein in synaptic terminals, wher e it colocalizes with synaptojanin. In vitro binding assays demonstrat e that endophilin binds through its src homology 3 domain to synaptoja nin, and immunoprecipitation analysis with the antiendophilin antibody reveals that endophilin is stably associated with synaptojanin in the nerve terminal. Immunoprecipitation with an antibody against amphiphy sin 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 synaptoja nin 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 fu nction.