N-glycosylation is essential for vesicular targeting of synaptotagmin 1.
Synaptotagmins 1 and 7 are candidate Ca(2+) sensors for exocytosis localized to synaptic vesicles and plasma membranes, respectively. We now show that the N-terminal intraluminal sequence of synaptotagmin 1, when transplanted onto synaptotagmin 7, redirects synaptotagmin 7 from the plasma membrane to secretory vesicles. Conversely, mutation of the N-terminal N-glycosylation site of synaptotagmin 1 redirects synaptotagmin 1 from vesicles to the plasma membrane. In cultured hippocampal neurons, the plasma membrane-localized mutant of synaptotagmin 1 suppressed the readily releasable pool of synaptic vesicles, whereas wild-type synaptotagmin 1 did not. In addition to the intraluminal N-glycosylation site, the cytoplasmic C(2) domains of synaptotagmin 1 were required for correct targeting but could be functionally replaced by the C(2) domains of synaptotagmin 7. Our data suggest that the intravesicular N-glycosylation site of synaptotagmin 1 collaborates with its cytoplasmic C(2) domains in directing synaptotagmin 1 to synaptic vesicles via a novel N-glycosylation-dependent mechanism.
Han
Weiping
W
Center for Basic Neuroscience, Department of Molecular Genetics, The University of Texas Southwestern Medical Center, Dallas, 75390, USA.
Rhee
Jeong-Seop
JS
Maximov
Anton
A
Lao
Ye
Y
Mashimo
Tomoyuki
T
Rosenmund
Christian
C
Südhof
Thomas C
TC
D000375:Aging / Q000378:metabolism; D000818:Animals; D001692:Biological Transport / Q000235:genetics; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism; D005822:Genetic Vectors; D006031:Glycosylation; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation / Q000502:physiology; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D009490:Neurosecretory Systems / Q000166:cytology / Q000378:metabolism; D018377:Neurotransmitter Agents / Q000378:metabolism; D016716:PC12 Cells; D017434:Protein Structure, Tertiary; D051381:Rats; D012672:Semliki forest virus / Q000235:genetics; D013572:Synaptic Vesicles / Q000502:physiology*; D050863:Synaptotagmin I; D050857:Synaptotagmins; D014162:Transfection
D002135:Calcium-Binding Proteins; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D018377:Neurotransmitter Agents; D050863:Synaptotagmin I; C494311:Syt1 protein, mouse; C494312:Syt1 protein, rat; D050857:Synaptotagmins