Identification of SNAREs involved in synaptotagmin VII-regulated lysosomal exocytosis.
Ca2+-regulated exocytosis of lysosomes has been recognized recently as a ubiquitous process, important for the repair of plasma membrane wounds. Lysosomal exocytosis is regulated by synaptotagmin VII, a member of the synaptotagmin family of Ca2+-binding proteins localized on lysosomes. Here we show that Ca2+-dependent interaction of the synaptotagmin VII C(2)A domain with SNAP-23 is facilitated by syntaxin 4. Specific interactions also occurred in cell lysates between the plasma membrane t-SNAREs SNAP-23 and syntaxin 4 and the lysosomal v-SNARE TI-VAMP/VAMP7. Following cytosolic Ca2+ elevation, SDS-resistant complexes containing SNAP-23, syntaxin 4, and TI-VAMP/VAMP7 were detected on membrane fractions. Lysosomal exocytosis was inhibited by the SNARE domains of syntaxin 4 and TI-VAMP/VAMP7 and by cleavage of SNAP-23 with botulinum neurotoxin E, thereby functionally implicating these SNAREs in Ca2+-regulated exocytosis of conventional lysosomes.
Rao
Swathi K
SK
Section of Microbial Pathogenesis and Department of Cell Biology, Yale University School of Medicine, Boyer Center for Molecular Medicine, 295 Congress Avenue, New Haven, CT 06511, USA.
Huynh
Chau
C
Proux-Gillardeaux
Veronique
V
Galli
Thierry
T
Andrews
Norma W
NW
D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D001426:Bacterial Proteins; D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D004247:DNA / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D018274:Electroporation; D005089:Exocytosis; D005434:Flow Cytometry; D051907:Lysosomal Membrane Proteins; D008247:Lysosomes / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism / Q000502:physiology*; D008565:Membrane Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D009419:Nerve Tissue Proteins / Q000378:metabolism / Q000502:physiology*; D011233:Precipitin Tests; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D050765:Qa-SNARE Proteins; D050766:Qb-SNARE Proteins; D050767:Qc-SNARE Proteins; D050683:R-SNARE Proteins; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D050600:SNARE Proteins; D013301:Streptolysins / Q000494:pharmacology; D050857:Synaptotagmins; D033921:Vesicular Transport Proteins; D001619:beta-N-Acetylhexosaminidases / Q000737:chemistry
D015703:Antigens, CD; D001426:Bacterial Proteins; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D051907:Lysosomal Membrane Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; D050765:Qa-SNARE Proteins; D050766:Qb-SNARE Proteins; D050767:Qc-SNARE Proteins; D050683:R-SNARE Proteins; D011994:Recombinant Proteins; C100493:SNAP23 protein, human; D050600:SNARE Proteins; D013301:Streptolysins; C494304:Syt7 protein, rat; C496926:Vamp7 protein, rat; D033921:Vesicular Transport Proteins; C025964:streptolysin O; D050857:Synaptotagmins; D004247:DNA; D001619:beta-N-Acetylhexosaminidases; D002118:Calcium