Synaptotagmin VII regulates Ca2+-dependent exocytosis of lysosomes in fibroblasts

Synaptotagmins (Syts) are transmembrane proteins with two Ca2+-binding C-2 domains in their cytosolic region. Syt I, the most widely studied isoform, has been proposed to function as a Ca2+ 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, Rite. 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 Ca2+ concentrations ([Ca2+]) below 10 mu M, whereas other isoforms require 200-500 mu M [Ca2+] or show no Ca2+-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 mu M [Ca2+] (Rodriguez, 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 Ca2+-triggered secretion of P-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 C2A domain are also inhibitory in both assays, indicating that Syt VII plays a key role in the regulation of Ca2+-dependent lysosome exocytosis.