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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

Exocytosis is uncoupled from RAB-3 synaptic vesicle dissociation in snt-1 mutants.(A) The failure of RAB-3/SV dissociation caused by exocytosis mutants, including unc-2 and unc-13, is bypassed by mutation of snt-1. Yellow arrows indicate the cell bodies along the ventral cord. Scale bar, 5 µm. (B) Quantification of the synaptic enrichment in the genotypes shown in (A). Data are presented as mean ± SD. *p < 0.05; NS, not significant.DOI:http://dx.doi.org/10.7554/eLife.05118.012
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fig7s1: Exocytosis is uncoupled from RAB-3 synaptic vesicle dissociation in snt-1 mutants.(A) The failure of RAB-3/SV dissociation caused by exocytosis mutants, including unc-2 and unc-13, is bypassed by mutation of snt-1. Yellow arrows indicate the cell bodies along the ventral cord. Scale bar, 5 µm. (B) Quantification of the synaptic enrichment in the genotypes shown in (A). Data are presented as mean ± SD. *p < 0.05; NS, not significant.DOI:http://dx.doi.org/10.7554/eLife.05118.012

Mentions: Could the enhanced GFP:RAB-3 signal in the puncta indeed reflect the failure of exocytosis? Previous studies suggest that RAB3 dissociation from SV membranes is inhibited when SV exocytosis is disrupted by blocking either Ca2+ influx or membrane fusion (Fischer von Mollard et al., 1991; Fischer von Mollard et al., 1994; Stahl et al., 1996). We examined two exocytosis mutants, unc-2 and unc-13. unc-2 encodes the alpha subunit of the voltage-gated Ca2+ channel (Schafer and Kenyon, 1995). In unc-2 mutants, we found that the GFP signal is significantly enriched within the punctate regions and the GFP::RAB-3 puncta are larger and brighter compared to wild type (Figure 7—figure supplement 1A,B). unc-13 encodes the Munc13 homolog in worms and loss of function of unc-13 results in blockage of membrane fusion during SV exocytosis (Aravamudan et al., 1999; Richmond et al., 1999; Varoqueaux et al., 2002). We found that the GFP signal was enhanced in the punctate regions in unc-13 mutants, as in unc-2 mutants (Figure 7—figure supplement 1A,B). Blocking SV exocytosis resulted in failure of RAB-3 to dissociate from SVs; thus the enlarged GFP::RAB-3 puncta indeed indicate the decreased dissociation of RAB-3 protein from SVs. Taken together, the data above indicate that the Ca2+-binding capability is essential for SNT-1 function in mediating the RAB-3 SV dissociation induced by Ca2+-triggered exocytosis.


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

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

Exocytosis is uncoupled from RAB-3 synaptic vesicle dissociation in snt-1 mutants.(A) The failure of RAB-3/SV dissociation caused by exocytosis mutants, including unc-2 and unc-13, is bypassed by mutation of snt-1. Yellow arrows indicate the cell bodies along the ventral cord. Scale bar, 5 µm. (B) Quantification of the synaptic enrichment in the genotypes shown in (A). Data are presented as mean ± SD. *p < 0.05; NS, not significant.DOI:http://dx.doi.org/10.7554/eLife.05118.012
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4374511&req=5

fig7s1: Exocytosis is uncoupled from RAB-3 synaptic vesicle dissociation in snt-1 mutants.(A) The failure of RAB-3/SV dissociation caused by exocytosis mutants, including unc-2 and unc-13, is bypassed by mutation of snt-1. Yellow arrows indicate the cell bodies along the ventral cord. Scale bar, 5 µm. (B) Quantification of the synaptic enrichment in the genotypes shown in (A). Data are presented as mean ± SD. *p < 0.05; NS, not significant.DOI:http://dx.doi.org/10.7554/eLife.05118.012
Mentions: Could the enhanced GFP:RAB-3 signal in the puncta indeed reflect the failure of exocytosis? Previous studies suggest that RAB3 dissociation from SV membranes is inhibited when SV exocytosis is disrupted by blocking either Ca2+ influx or membrane fusion (Fischer von Mollard et al., 1991; Fischer von Mollard et al., 1994; Stahl et al., 1996). We examined two exocytosis mutants, unc-2 and unc-13. unc-2 encodes the alpha subunit of the voltage-gated Ca2+ channel (Schafer and Kenyon, 1995). In unc-2 mutants, we found that the GFP signal is significantly enriched within the punctate regions and the GFP::RAB-3 puncta are larger and brighter compared to wild type (Figure 7—figure supplement 1A,B). unc-13 encodes the Munc13 homolog in worms and loss of function of unc-13 results in blockage of membrane fusion during SV exocytosis (Aravamudan et al., 1999; Richmond et al., 1999; Varoqueaux et al., 2002). We found that the GFP signal was enhanced in the punctate regions in unc-13 mutants, as in unc-2 mutants (Figure 7—figure supplement 1A,B). Blocking SV exocytosis resulted in failure of RAB-3 to dissociate from SVs; thus the enlarged GFP::RAB-3 puncta indeed indicate the decreased dissociation of RAB-3 protein from SVs. Taken together, the data above indicate that the Ca2+-binding capability is essential for SNT-1 function in mediating the RAB-3 SV dissociation induced by Ca2+-triggered exocytosis.

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