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Synaptotagmin 1 directs repetitive release by coupling vesicle exocytosis to the Rab3 cycle.

Cheng Y, Wang J, Wang Y, Ding M - Elife (2015)

Bottom Line: How this harmonization is achieved is not known.In the absence of Ca(2+), synaptotagmin 1 binds to Rab3 GTPase activating protein (GAP) and inhibits the GTP hydrolysis of Rab3 protein.In the presence of Ca(2+), synaptotagmin 1 releases Rab3 GAP and promotes membrane disassociation of Rab3.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.

ABSTRACT
In response to Ca(2+) influx, a synapse needs to release neurotransmitters quickly while immediately preparing for repeat firing. How this harmonization is achieved is not known. In this study, we found that the Ca(2+) sensor synaptotagmin 1 orchestrates the membrane association/disassociation cycle of Rab3, which functions in activity-dependent recruitment of synaptic vesicles. In the absence of Ca(2+), synaptotagmin 1 binds to Rab3 GTPase activating protein (GAP) and inhibits the GTP hydrolysis of Rab3 protein. Rab3 GAP resides on synaptic vesicles, and synaptotagmin 1 is essential for the synaptic localization of Rab3 GAP. In the presence of Ca(2+), synaptotagmin 1 releases Rab3 GAP and promotes membrane disassociation of Rab3. Without synaptotagmin 1, the tight coupling between vesicle exocytosis and Rab3 membrane disassociation is disrupted. We uncovered the long-sought molecular apparatus linking vesicle exocytosis to Rab3 cycling and we also revealed the important function of synaptotagmin 1 in repetitive synaptic vesicle release.

No MeSH data available.


Related in: MedlinePlus

RBG-1 associates with the C2B domain of SNT-1.(A) RBG-1 is precipitated by the intracellular domain (C2AB) of SNT-1. (B) The SNT-1 intracellular domain is precipitated by RBG-1. (C) The C2B domain of SNT-1 binds to RBG-1. (D) SNT-1 without the C2B domain fails to rescue the snt-1 mutant phenotype. Yellow arrows indicate cell bodies. The schematic diagram shows the transmembrane (TM) and intracellular calcium-binding domains (C2A and C2B) of SNT-1. Scale bar, 5 µm.DOI:http://dx.doi.org/10.7554/eLife.05118.010
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fig6: RBG-1 associates with the C2B domain of SNT-1.(A) RBG-1 is precipitated by the intracellular domain (C2AB) of SNT-1. (B) The SNT-1 intracellular domain is precipitated by RBG-1. (C) The C2B domain of SNT-1 binds to RBG-1. (D) SNT-1 without the C2B domain fails to rescue the snt-1 mutant phenotype. Yellow arrows indicate cell bodies. The schematic diagram shows the transmembrane (TM) and intracellular calcium-binding domains (C2A and C2B) of SNT-1. Scale bar, 5 µm.DOI:http://dx.doi.org/10.7554/eLife.05118.010

Mentions: The SNT-1-dependent SV association of RBG-1 suggests a direct association between SNT-1 and RBG-1. We co-expressed full-length RBG-1 and the cytosolic domain of SNT-1 (C2AB) in HEK293FT cells. After affinity purification, the RBG-1 protein was incubated with the SNT-1 C2AB fragment. In contrast to the mock-transfected sample, RBG-1 was effectively co-precipitated with the cytosolic region of SNT-1 (Figure 6A). The SNT-1 cytosolic portion was also co-precipitated by RBG-1 (Figure 6B). We next asked which domain of the SNT-1 cytosolic region is required for this binding. The cytosolic region of SNT-1 contains a C2A and a C2B motif. When the C2A domain was deleted, the remaining C2B motif retained the RBG-1 binding activity (Figure 6C). In contrast, when C2B was removed, the C2A domain alone could not bind to RBG-1 (Figure 6C). Therefore, the C2B domain is required for SNT-1 binding to RBG-1.10.7554/eLife.05118.010Figure 6.RBG-1 associates with the C2B domain of SNT-1.


Synaptotagmin 1 directs repetitive release by coupling vesicle exocytosis to the Rab3 cycle.

Cheng Y, Wang J, Wang Y, Ding M - Elife (2015)

RBG-1 associates with the C2B domain of SNT-1.(A) RBG-1 is precipitated by the intracellular domain (C2AB) of SNT-1. (B) The SNT-1 intracellular domain is precipitated by RBG-1. (C) The C2B domain of SNT-1 binds to RBG-1. (D) SNT-1 without the C2B domain fails to rescue the snt-1 mutant phenotype. Yellow arrows indicate cell bodies. The schematic diagram shows the transmembrane (TM) and intracellular calcium-binding domains (C2A and C2B) of SNT-1. Scale bar, 5 µm.DOI:http://dx.doi.org/10.7554/eLife.05118.010
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4374511&req=5

fig6: RBG-1 associates with the C2B domain of SNT-1.(A) RBG-1 is precipitated by the intracellular domain (C2AB) of SNT-1. (B) The SNT-1 intracellular domain is precipitated by RBG-1. (C) The C2B domain of SNT-1 binds to RBG-1. (D) SNT-1 without the C2B domain fails to rescue the snt-1 mutant phenotype. Yellow arrows indicate cell bodies. The schematic diagram shows the transmembrane (TM) and intracellular calcium-binding domains (C2A and C2B) of SNT-1. Scale bar, 5 µm.DOI:http://dx.doi.org/10.7554/eLife.05118.010
Mentions: The SNT-1-dependent SV association of RBG-1 suggests a direct association between SNT-1 and RBG-1. We co-expressed full-length RBG-1 and the cytosolic domain of SNT-1 (C2AB) in HEK293FT cells. After affinity purification, the RBG-1 protein was incubated with the SNT-1 C2AB fragment. In contrast to the mock-transfected sample, RBG-1 was effectively co-precipitated with the cytosolic region of SNT-1 (Figure 6A). The SNT-1 cytosolic portion was also co-precipitated by RBG-1 (Figure 6B). We next asked which domain of the SNT-1 cytosolic region is required for this binding. The cytosolic region of SNT-1 contains a C2A and a C2B motif. When the C2A domain was deleted, the remaining C2B motif retained the RBG-1 binding activity (Figure 6C). In contrast, when C2B was removed, the C2A domain alone could not bind to RBG-1 (Figure 6C). Therefore, the C2B domain is required for SNT-1 binding to RBG-1.10.7554/eLife.05118.010Figure 6.RBG-1 associates with the C2B domain of SNT-1.

Bottom Line: How this harmonization is achieved is not known.In the absence of Ca(2+), synaptotagmin 1 binds to Rab3 GTPase activating protein (GAP) and inhibits the GTP hydrolysis of Rab3 protein.In the presence of Ca(2+), synaptotagmin 1 releases Rab3 GAP and promotes membrane disassociation of Rab3.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.

ABSTRACT
In response to Ca(2+) influx, a synapse needs to release neurotransmitters quickly while immediately preparing for repeat firing. How this harmonization is achieved is not known. In this study, we found that the Ca(2+) sensor synaptotagmin 1 orchestrates the membrane association/disassociation cycle of Rab3, which functions in activity-dependent recruitment of synaptic vesicles. In the absence of Ca(2+), synaptotagmin 1 binds to Rab3 GTPase activating protein (GAP) and inhibits the GTP hydrolysis of Rab3 protein. Rab3 GAP resides on synaptic vesicles, and synaptotagmin 1 is essential for the synaptic localization of Rab3 GAP. In the presence of Ca(2+), synaptotagmin 1 releases Rab3 GAP and promotes membrane disassociation of Rab3. Without synaptotagmin 1, the tight coupling between vesicle exocytosis and Rab3 membrane disassociation is disrupted. We uncovered the long-sought molecular apparatus linking vesicle exocytosis to Rab3 cycling and we also revealed the important function of synaptotagmin 1 in repetitive synaptic vesicle release.

No MeSH data available.


Related in: MedlinePlus