title: Discrete residues in the c(2)b domain of synaptotagmin I independently specify endocytic rate and synaptic vesicle size.
abstract: It has been demonstrated that synapses lacking functional synaptotagmin I (Syt I) have a decreased rate of synaptic vesicle endocytosis. Beyond this, the function of Syt I during endocytosis remains undefined. Here, we demonstrate that a decreased rate of endocytosis in syt(null) mutants correlates with a stimulus-dependent perturbation of membrane internalization, assayed ultrastructurally. We then separate the mechanisms that control endocytic rate and vesicle size by mapping these processes to discrete residues in the Syt I C(2)B domain. Mutation of a poly-lysine motif alters vesicle size but not endocytic rate, whereas the mutation of calcium-coordinating aspartate residues (syt-D3,4N) alters endocytic rate but not vesicle size. Finally, slowed endocytic rate in the syt-D3,4N animals, but not syt(null) animals, can be rescued by elevating extracellular calcium concentration, supporting the conclusion that calcium coordination within the C(2)B domain contributes to the control of endocytic rate.
authors.affiliation: Department of Biochemistry and Biophysics, University of California, San Francisco, Rock Hall, 1550 Fourth Street, Box 2822, San Francisco, California 94158, USA.
authors.forename: Kira E
authors.initials: KE
authors.lastname: Poskanzer
authors.forename: Richard D
authors.initials: RD
authors.lastname: Fetter
authors.forename: Graeme W
authors.initials: GW
authors.lastname: Davis
mesh_terms: D033965:Adaptor Protein Complex alpha Subunits / Q000378:metabolism; D000818:Animals; D030801:Animals, Genetically Modified; D001224:Aspartic Acid / Q000235:genetics / Q000378:metabolism; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004307:Dose-Response Relationship, Radiation; D004330:Drosophila; D029721:Drosophila Proteins / Q000378:metabolism*; D004558:Electric Stimulation / Q000379:methods; D004705:Endocytosis / Q000235:genetics / Q000502:physiology*; D019706:Excitatory Postsynaptic Potentials / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects; D049452:Green Fluorescent Proteins / Q000494:pharmacology; D007150:Immunohistochemistry / Q000379:methods; D007814:Larva; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D008564:Membrane Potentials / Q000502:physiology; D046529:Microscopy, Electron, Transmission / Q000379:methods; D009154:Mutation; D010775:Photic Stimulation / Q000379:methods; D017434:Protein Structure, Tertiary; D013569:Synapses / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D013572:Synaptic Vesicles / Q000502:physiology* / Q000648:ultrastructure; D050863:Synaptotagmin I / Q000737:chemistry / Q000378:metabolism*; D013997:Time Factors
chemical_list: D033965:Adaptor Protein Complex alpha Subunits; D002135:Calcium-Binding Proteins; D029721:Drosophila Proteins; D050863:Synaptotagmin I; D049452:Green Fluorescent Proteins; D001224:Aspartic Acid; D008239:Lysine; D002118:Calcium