title: Synaptotagmin VII regulates Ca(2+)-dependent exocytosis of lysosomes in fibroblasts.
abstract: Synaptotagmins (Syts) are transmembrane proteins with two Ca(2+)-binding C(2) domains in their cytosolic region. Syt I, the most widely studied isoform, has been proposed to function as a Ca(2+) sensor in synaptic vesicle exocytosis. Several of the twelve known Syts are expressed primarily in brain, while a few are ubiquitous (Sudhof, T.C., and J. Rizo. 1996. Neuron. 17: 379-388; Butz, S., R. Fernandez-Chacon, F. Schmitz, R. Jahn, and T.C. Sudhof. 1999. J. Biol. Chem. 274:18290-18296). The ubiquitously expressed Syt VII binds syntaxin at free Ca(2+) concentrations ([Ca(2+)]) below 10 microM, whereas other isoforms require 200-500 microM [Ca(2+)] or show no Ca(2+)-dependent syntaxin binding (Li, C., B. Ullrich, Z. Zhang, R.G.W. Anderson, N. Brose, and T.C. Sudhof. 1995. Nature. 375:594-599). We investigated the involvement of Syt VII in the exocytosis of lysosomes, which is triggered in several cell types at 1-5 microM [Ca(2+)] (Rodríguez, A., P. Webster, J. Ortego, and N.W. Andrews. 1997. J. Cell Biol. 137:93-104). Here, we show that Syt VII is localized on dense lysosomes in normal rat kidney (NRK) fibroblasts, and that GFP-tagged Syt VII is targeted to lysosomes after transfection. Recombinant fragments containing the C(2)A domain of Syt VII inhibit Ca(2+)-triggered secretion of beta-hexosaminidase and surface translocation of Lgp120, whereas the C(2)A domain of the neuronal- specific isoform, Syt I, has no effect. Antibodies against the Syt VII C(2)A domain are also inhibitory in both assays, indicating that Syt VII plays a key role in the regulation of Ca(2+)-dependent lysosome exocytosis.
authors.affiliation: Section of Microbial Pathogenesis, Boyer Center for Molecular Medicine, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
authors.forename: I
authors.initials: I
authors.lastname: Martinez
authors.forename: S
authors.initials: S
authors.lastname: Chakrabarti
authors.forename: T
authors.initials: T
authors.lastname: Hellevik
authors.forename: J
authors.initials: J
authors.lastname: Morehead
authors.forename: K
authors.initials: K
authors.lastname: Fowler
authors.forename: N W
authors.initials: NW
authors.lastname: Andrews
mesh_terms: D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000502:physiology*; D002118:Calcium / Q000502:physiology*; D002135:Calcium-Binding Proteins / Q000378:metabolism; D002460:Cell Line; D002463:Cell Membrane Permeability; D003001:Cloning, Molecular; D005089:Exocytosis / Q000502:physiology*; D005347:Fibroblasts / Q000502:physiology / Q000648:ultrastructure; D007668:Kidney; D007700:Kinetics; D008247:Lysosomes / Q000502:physiology* / Q000648:ultrastructure; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D020033:Protein Isoforms / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D050857:Synaptotagmins; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism
chemical_list: D002135:Calcium-Binding Proteins; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; D011994:Recombinant Proteins; C494304:Syt7 protein, rat; D050857:Synaptotagmins; D001619:beta-N-Acetylhexosaminidases; D002118:Calcium