Synaptojanin forms two separate complexes in the nerve terminal. Interactions with endophilin and amphiphysin.
Endophilin is a recently discovered src homology 3 domain-containing protein that is a major in vitro binding 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.
Micheva
K D
KD
Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, 3801 Rue University, Montreal, Quebec H3A 2B4, Canada.
Kay
B K
BK
McPherson
P S
PS
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism*; D002352:Carrier Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D002458:Cell Fractionation; D002846:Chromatography, Affinity; D004791:Enzyme Inhibitors / Q000378:metabolism; D007150:Immunohistochemistry; D008297:Male; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D009928:Organ Specificity; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology; D010739:Phospholipase D / Q000037:antagonists & inhibitors; D010744:Phosphoric Monoester Hydrolases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D017729:Presynaptic Terminals / Q000378:metabolism* / Q000648:ultrastructure; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000096:biosynthesis; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D000906:Antibodies; D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C075637:amphiphysin; C097702:synaptojanin; D010744:Phosphoric Monoester Hydrolases; D010739:Phospholipase D