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The Sec1p/Munc18 (SM) protein, Vps45p, cycles on and off membranes during vesicle transport.

Bryant NJ, James DE - J. Cell Biol. (2003)

Bottom Line: Here, we show that glc7 mutant cells accumulate SNARE complexes.These complexes are clearly different from those found in either wild-type or sec18-1 cells as the Sec1p/Munc18 (SM) protein Vps45p does not bind to them.Vps45p dissociates from the membrane in the absence of PP1 activity, but rapidly reassociates after its reactivation.

View Article: PubMed Central - PubMed

Affiliation: The Garvan Institute of Medical Research, Darlinghurst, Sydney, NSW, Australia 2010. n.bryant@bio.gla.ac.uk

ABSTRACT
Protein phosphatase 1 (PP1, Glc7p) functions in the final stage of SNARE-mediated vesicle transport between docking and fusion. During this process, trans-SNARE complexes, formed between molecules in opposing membranes, convert to cis-complexes, with all participants in the same lipid bilayer. Here, we show that glc7 mutant cells accumulate SNARE complexes. These complexes are clearly different from those found in either wild-type or sec18-1 cells as the Sec1p/Munc18 (SM) protein Vps45p does not bind to them. Given that PP1 controls fusion, the SNARE complexes that accumulate in glc7 mutants likely represent trans-SNARE complexes. Vps45p dissociates from the membrane in the absence of PP1 activity, but rapidly reassociates after its reactivation. These data reveal that SM proteins cycle on and off membranes in a stage-specific manner during the vesicle transport reaction, and suggest that protein phosphorylation plays a key role in the regulation of this cycle.

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Model for the interaction of Vps45p with Tlg2p during SNARE complex assembly–disassembly. Model depicting the association and dissociation of Vps45p with membranes at different stages of the SNARE assembly–disassembly cycle. The cycle begins with the t-SNARE/SM heterodimer (Tlg2p/Vps45p), which precedes the docking reaction involving the Rab protein Vps21p. Docking results in the formation of a trans-SNARE complex involving the v-SNARE on the transport vesicle and the t-SNAREs in the target membrane. The SNARE complex is found in a cis-configuration following merging of the two lipid bilayers during the fusion reaction (involving the protein phosphatase, Glc7p), and is subsequently disassembled by the ATPase, Sec18p. Vps45p dissociates from membranes either before or during formation of the trans-complex in concert with the action of the Rab protein and/or the phosphatase and then rebinds to the cis-SNARE complex.
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fig5: Model for the interaction of Vps45p with Tlg2p during SNARE complex assembly–disassembly. Model depicting the association and dissociation of Vps45p with membranes at different stages of the SNARE assembly–disassembly cycle. The cycle begins with the t-SNARE/SM heterodimer (Tlg2p/Vps45p), which precedes the docking reaction involving the Rab protein Vps21p. Docking results in the formation of a trans-SNARE complex involving the v-SNARE on the transport vesicle and the t-SNAREs in the target membrane. The SNARE complex is found in a cis-configuration following merging of the two lipid bilayers during the fusion reaction (involving the protein phosphatase, Glc7p), and is subsequently disassembled by the ATPase, Sec18p. Vps45p dissociates from membranes either before or during formation of the trans-complex in concert with the action of the Rab protein and/or the phosphatase and then rebinds to the cis-SNARE complex.

Mentions: The SNARE assembly cycle depicted in Fig. 5 illustrates the dynamic equilibrium between the t-SNARE Tlg2p in its monomeric state, and as part of trans- and cis-complexes. Each one of these states defines an intermediate in the vesicle docking–fusion process. We have demonstrated that Vps45p binds to Tlg2p in this state before the formation of the trans-SNARE complex in a Rab (Vps21p)-dependent process (Lupashin and Waters, 1997; Grote and Novick, 1999). We show that Vps45p dissociates at this stage of the cycle and is released into the cytosol. The conversion of trans-SNARE complexes to the cis-configuration defines membrane fusion (Ungermann et al., 1998), and also represents the starting point for another round of vesicle transport. It is here that Vps45p reenters the cycle by binding to the cis-complex. The cycle is completed after disassembly of cis-SNARE complexes with Vps45p bound by Sec18p presumably resulting in the Vps45p–Tlg2p complex that is then ready to receive incoming vesicles in the process of docking.


The Sec1p/Munc18 (SM) protein, Vps45p, cycles on and off membranes during vesicle transport.

Bryant NJ, James DE - J. Cell Biol. (2003)

Model for the interaction of Vps45p with Tlg2p during SNARE complex assembly–disassembly. Model depicting the association and dissociation of Vps45p with membranes at different stages of the SNARE assembly–disassembly cycle. The cycle begins with the t-SNARE/SM heterodimer (Tlg2p/Vps45p), which precedes the docking reaction involving the Rab protein Vps21p. Docking results in the formation of a trans-SNARE complex involving the v-SNARE on the transport vesicle and the t-SNAREs in the target membrane. The SNARE complex is found in a cis-configuration following merging of the two lipid bilayers during the fusion reaction (involving the protein phosphatase, Glc7p), and is subsequently disassembled by the ATPase, Sec18p. Vps45p dissociates from membranes either before or during formation of the trans-complex in concert with the action of the Rab protein and/or the phosphatase and then rebinds to the cis-SNARE complex.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199362&req=5

fig5: Model for the interaction of Vps45p with Tlg2p during SNARE complex assembly–disassembly. Model depicting the association and dissociation of Vps45p with membranes at different stages of the SNARE assembly–disassembly cycle. The cycle begins with the t-SNARE/SM heterodimer (Tlg2p/Vps45p), which precedes the docking reaction involving the Rab protein Vps21p. Docking results in the formation of a trans-SNARE complex involving the v-SNARE on the transport vesicle and the t-SNAREs in the target membrane. The SNARE complex is found in a cis-configuration following merging of the two lipid bilayers during the fusion reaction (involving the protein phosphatase, Glc7p), and is subsequently disassembled by the ATPase, Sec18p. Vps45p dissociates from membranes either before or during formation of the trans-complex in concert with the action of the Rab protein and/or the phosphatase and then rebinds to the cis-SNARE complex.
Mentions: The SNARE assembly cycle depicted in Fig. 5 illustrates the dynamic equilibrium between the t-SNARE Tlg2p in its monomeric state, and as part of trans- and cis-complexes. Each one of these states defines an intermediate in the vesicle docking–fusion process. We have demonstrated that Vps45p binds to Tlg2p in this state before the formation of the trans-SNARE complex in a Rab (Vps21p)-dependent process (Lupashin and Waters, 1997; Grote and Novick, 1999). We show that Vps45p dissociates at this stage of the cycle and is released into the cytosol. The conversion of trans-SNARE complexes to the cis-configuration defines membrane fusion (Ungermann et al., 1998), and also represents the starting point for another round of vesicle transport. It is here that Vps45p reenters the cycle by binding to the cis-complex. The cycle is completed after disassembly of cis-SNARE complexes with Vps45p bound by Sec18p presumably resulting in the Vps45p–Tlg2p complex that is then ready to receive incoming vesicles in the process of docking.

Bottom Line: Here, we show that glc7 mutant cells accumulate SNARE complexes.These complexes are clearly different from those found in either wild-type or sec18-1 cells as the Sec1p/Munc18 (SM) protein Vps45p does not bind to them.Vps45p dissociates from the membrane in the absence of PP1 activity, but rapidly reassociates after its reactivation.

View Article: PubMed Central - PubMed

Affiliation: The Garvan Institute of Medical Research, Darlinghurst, Sydney, NSW, Australia 2010. n.bryant@bio.gla.ac.uk

ABSTRACT
Protein phosphatase 1 (PP1, Glc7p) functions in the final stage of SNARE-mediated vesicle transport between docking and fusion. During this process, trans-SNARE complexes, formed between molecules in opposing membranes, convert to cis-complexes, with all participants in the same lipid bilayer. Here, we show that glc7 mutant cells accumulate SNARE complexes. These complexes are clearly different from those found in either wild-type or sec18-1 cells as the Sec1p/Munc18 (SM) protein Vps45p does not bind to them. Given that PP1 controls fusion, the SNARE complexes that accumulate in glc7 mutants likely represent trans-SNARE complexes. Vps45p dissociates from the membrane in the absence of PP1 activity, but rapidly reassociates after its reactivation. These data reveal that SM proteins cycle on and off membranes in a stage-specific manner during the vesicle transport reaction, and suggest that protein phosphorylation plays a key role in the regulation of this cycle.

Show MeSH
Related in: MedlinePlus