Dual roles of the C2B domain of synaptotagmin I in synchronizing Ca2+-dependent neurotransmitter release

Tei-ichi Nishiki, George J. Augustine

Research output: Contribution to journalArticle

98 Citations (Scopus)

Abstract

Although the vesicular protein synaptotagmin I contains two Ca 2+-binding domains (C2A and C2B), Ca 2+ binding to the C2B domain is more important for triggering synchronous neurotransmitter release. We have used point mutagenesis to determine the functional contributions of the five negatively charged aspartate (Asp) residues that constitute the Ca2+-binding sites in the C2B domain of synaptotagmin I. Transfecting wild-type synaptotagmin I DNA into cultured hippocampal neurons from synaptotagmin I knock-out mice rescued Ca2+-dependent synchronous transmitter release and reduced a slower, asynchronous component of release, indicating that synaptotagmin I suppresses asynchronous release. Mutating either the second or third Asp residues of the C2B domain potently inhibited the ability of synaptotagmin I to rescue synchronous release but did not change its ability to suppress asynchronous release. Synaptotagmin I with mutations in the first or fourth Asp residues of the C2B domain partially rescued synchronous release and partially suppressed asynchronous release, whereas neutralizing the fifth Asp residue had no effect on the ability of synaptotagmin I to rescue transmitter release. Thus, we conclude that the C2B domain of synaptotagmin I regulates neurotransmitter release in at least two ways. Synchronous release absolutely requires binding of Ca2+ to the second and third Asp residues in this domain. For the suppression of asynchronous release, Ca2+ binding to the C2B domain of synaptotagmin I apparently is not necessary because mutation of the second Asp residue inhibits Ca2+ binding, yet still allows this protein to suppress asynchronous release.

Original languageEnglish
Pages (from-to)8542-8550
Number of pages9
JournalJournal of Neuroscience
Volume24
Issue number39
DOIs
Publication statusPublished - Sep 29 2004
Externally publishedYes

Fingerprint

Synaptotagmin I
Neurotransmitter Agents
Aspartic Acid
Mutation
Knockout Mice
Mutagenesis
Proteins

Keywords

  • Calcium
  • Exocytosis
  • Hippocampal neurons
  • SNARE
  • Synaptic transmission
  • Synaptic vesicle

ASJC Scopus subject areas

  • Neuroscience(all)

Cite this

Dual roles of the C2B domain of synaptotagmin I in synchronizing Ca2+-dependent neurotransmitter release. / Nishiki, Tei-ichi; Augustine, George J.

In: Journal of Neuroscience, Vol. 24, No. 39, 29.09.2004, p. 8542-8550.

Research output: Contribution to journalArticle

@article{0986916426324436a377dff6308cec0a,
title = "Dual roles of the C2B domain of synaptotagmin I in synchronizing Ca2+-dependent neurotransmitter release",
abstract = "Although the vesicular protein synaptotagmin I contains two Ca 2+-binding domains (C2A and C2B), Ca 2+ binding to the C2B domain is more important for triggering synchronous neurotransmitter release. We have used point mutagenesis to determine the functional contributions of the five negatively charged aspartate (Asp) residues that constitute the Ca2+-binding sites in the C2B domain of synaptotagmin I. Transfecting wild-type synaptotagmin I DNA into cultured hippocampal neurons from synaptotagmin I knock-out mice rescued Ca2+-dependent synchronous transmitter release and reduced a slower, asynchronous component of release, indicating that synaptotagmin I suppresses asynchronous release. Mutating either the second or third Asp residues of the C2B domain potently inhibited the ability of synaptotagmin I to rescue synchronous release but did not change its ability to suppress asynchronous release. Synaptotagmin I with mutations in the first or fourth Asp residues of the C2B domain partially rescued synchronous release and partially suppressed asynchronous release, whereas neutralizing the fifth Asp residue had no effect on the ability of synaptotagmin I to rescue transmitter release. Thus, we conclude that the C2B domain of synaptotagmin I regulates neurotransmitter release in at least two ways. Synchronous release absolutely requires binding of Ca2+ to the second and third Asp residues in this domain. For the suppression of asynchronous release, Ca2+ binding to the C2B domain of synaptotagmin I apparently is not necessary because mutation of the second Asp residue inhibits Ca2+ binding, yet still allows this protein to suppress asynchronous release.",
keywords = "Calcium, Exocytosis, Hippocampal neurons, SNARE, Synaptic transmission, Synaptic vesicle",
author = "Tei-ichi Nishiki and Augustine, {George J.}",
year = "2004",
month = "9",
day = "29",
doi = "10.1523/JNEUROSCI.2545-04.2004",
language = "English",
volume = "24",
pages = "8542--8550",
journal = "Journal of Neuroscience",
issn = "0270-6474",
publisher = "Society for Neuroscience",
number = "39",

}

TY - JOUR

T1 - Dual roles of the C2B domain of synaptotagmin I in synchronizing Ca2+-dependent neurotransmitter release

AU - Nishiki, Tei-ichi

AU - Augustine, George J.

PY - 2004/9/29

Y1 - 2004/9/29

N2 - Although the vesicular protein synaptotagmin I contains two Ca 2+-binding domains (C2A and C2B), Ca 2+ binding to the C2B domain is more important for triggering synchronous neurotransmitter release. We have used point mutagenesis to determine the functional contributions of the five negatively charged aspartate (Asp) residues that constitute the Ca2+-binding sites in the C2B domain of synaptotagmin I. Transfecting wild-type synaptotagmin I DNA into cultured hippocampal neurons from synaptotagmin I knock-out mice rescued Ca2+-dependent synchronous transmitter release and reduced a slower, asynchronous component of release, indicating that synaptotagmin I suppresses asynchronous release. Mutating either the second or third Asp residues of the C2B domain potently inhibited the ability of synaptotagmin I to rescue synchronous release but did not change its ability to suppress asynchronous release. Synaptotagmin I with mutations in the first or fourth Asp residues of the C2B domain partially rescued synchronous release and partially suppressed asynchronous release, whereas neutralizing the fifth Asp residue had no effect on the ability of synaptotagmin I to rescue transmitter release. Thus, we conclude that the C2B domain of synaptotagmin I regulates neurotransmitter release in at least two ways. Synchronous release absolutely requires binding of Ca2+ to the second and third Asp residues in this domain. For the suppression of asynchronous release, Ca2+ binding to the C2B domain of synaptotagmin I apparently is not necessary because mutation of the second Asp residue inhibits Ca2+ binding, yet still allows this protein to suppress asynchronous release.

AB - Although the vesicular protein synaptotagmin I contains two Ca 2+-binding domains (C2A and C2B), Ca 2+ binding to the C2B domain is more important for triggering synchronous neurotransmitter release. We have used point mutagenesis to determine the functional contributions of the five negatively charged aspartate (Asp) residues that constitute the Ca2+-binding sites in the C2B domain of synaptotagmin I. Transfecting wild-type synaptotagmin I DNA into cultured hippocampal neurons from synaptotagmin I knock-out mice rescued Ca2+-dependent synchronous transmitter release and reduced a slower, asynchronous component of release, indicating that synaptotagmin I suppresses asynchronous release. Mutating either the second or third Asp residues of the C2B domain potently inhibited the ability of synaptotagmin I to rescue synchronous release but did not change its ability to suppress asynchronous release. Synaptotagmin I with mutations in the first or fourth Asp residues of the C2B domain partially rescued synchronous release and partially suppressed asynchronous release, whereas neutralizing the fifth Asp residue had no effect on the ability of synaptotagmin I to rescue transmitter release. Thus, we conclude that the C2B domain of synaptotagmin I regulates neurotransmitter release in at least two ways. Synchronous release absolutely requires binding of Ca2+ to the second and third Asp residues in this domain. For the suppression of asynchronous release, Ca2+ binding to the C2B domain of synaptotagmin I apparently is not necessary because mutation of the second Asp residue inhibits Ca2+ binding, yet still allows this protein to suppress asynchronous release.

KW - Calcium

KW - Exocytosis

KW - Hippocampal neurons

KW - SNARE

KW - Synaptic transmission

KW - Synaptic vesicle

UR - http://www.scopus.com/inward/record.url?scp=4744344106&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=4744344106&partnerID=8YFLogxK

U2 - 10.1523/JNEUROSCI.2545-04.2004

DO - 10.1523/JNEUROSCI.2545-04.2004

M3 - Article

VL - 24

SP - 8542

EP - 8550

JO - Journal of Neuroscience

JF - Journal of Neuroscience

SN - 0270-6474

IS - 39

ER -