An In Vitro Assay of Trans-SNARE Complex Formation During Yeast Vacuole Fusion Using Epitope Tag-Free SNAREs

2018 ◽  
pp. 277-288
Author(s):  
Youngsoo Jun
Keyword(s):  
Complex Formation ◽  
In Vitro Assay ◽  
Snare Complex ◽  
Epitope Tag ◽  
Vitro Assay ◽  
Yeast Vacuole ◽  
Vacuole Fusion ◽  
Snare Complex Formation

scholarly journals Copper blocks V‐ATPase activity and SNARE complex formation to inhibit yeast vacuole fusion

Traffic ◽  
2019 ◽  
Vol 20 (11) ◽  
pp. 841-850 ◽  
Author(s):  
Gregory E. Miner ◽  
Katherine D. Sullivan ◽  
Chi Zhang ◽  
Logan R. Hurst ◽  
Matthew L. Starr ◽  
...  
Keyword(s):  
Complex Formation ◽  
Atpase Activity ◽  
Snare Complex ◽  
Yeast Vacuole ◽  
Vacuole Fusion ◽  
Snare Complex Formation

Author response for "Copper blocks V‐ATPase activity and SNARE complex formation to inhibit yeast vacuole fusion"

2019 ◽  
Author(s):  
Gregory E. Miner ◽  
Katherine D. Sullivan ◽  
Chi Zhang ◽  
Logan R. Hurst ◽  
Matthew L. Starr ◽  
...  
Keyword(s):  
Complex Formation ◽  
Atpase Activity ◽  
Snare Complex ◽  
Author Response ◽  
Yeast Vacuole ◽  
Vacuole Fusion ◽  
Snare Complex Formation

Author response for "Copper blocks V‐ATPase activity and SNARE complex formation to inhibit yeast vacuole fusion"

2019 ◽  
Author(s):  
Gregory E. Miner ◽  
Katherine D. Sullivan ◽  
Chi Zhang ◽  
Logan R. Hurst ◽  
Matthew L. Starr ◽  
...  
Keyword(s):  
Complex Formation ◽  
Atpase Activity ◽  
Snare Complex ◽  
Author Response ◽  
Yeast Vacuole ◽  
Vacuole Fusion ◽  
Snare Complex Formation

scholarly journals Phosphatidylinositol 3,5-Bisphosphate Regulates Yeast Vacuole Fusion at the Transition between trans-SNARE Complex Formation and Hemifusion

2018 ◽  
Author(s):  
Gregory E. Miner ◽  
Katherine D. Sullivan ◽  
Annie Guo ◽  
Brandon C. Jones ◽  
Matthew L. Starr ◽  
...  
Keyword(s):  
Complex Formation ◽  
Membrane Fusion ◽  
Molecular Switch ◽  
Snare Complex ◽  
Negative Regulator ◽  
Cellular Functions ◽  
Hyperosmotic Shock ◽  
Yeast Vacuole ◽  
Vacuole Fusion ◽  
Snare Complex Formation

ABSTRACTPhosphoinositides (PIs) regulate myriad cellular functions including membrane fusion, as exemplified by the yeast vacuole, which uses various PIs at different stages of fusion. In light of this, the effect of phos-phatidylinositol 3,5-bisphosphate [PI(3,5)P2] on vacuole fusion remains unknown. PI(3,5)P2 is made by the PI3P 5-kinase Fab1/PIKfyve and has been characterized as a regulator of vacuole fission during hyperosmotic shock where it interacts with the TRP family Ca2+ channel Yvc1. Here we demonstrate that exogenously added dioctanoyl (C8) PI(3,5)P2 abolishes homotypic vacuole fusion. This effect was not linked to interactions with Yvc1, as fusion was equally affected using yvc1Δ vacuoles. Thus, the effects of C8-PI(3,5)P2 on fusion versus fission operate through distinct mechanisms. Further testing showed that C8-PI(3,5)P2 inhibited vacuole fusion after the formation of trans-SNARE pairs. Although SNARE complex formation was unaffected we found that C8-PI(3,5)P2 strongly inhibited hemifusion. Overproduction of endogenous PI(3,5)P2 by the fab1T2250A hyperactive kinase mutant also inhibited at the hemifusion stage, bolstering the model in which PI(3,5)P2 inhibits fusion when present elevated levels. Taken together, this work identifies a novel function for PI(3,5)P2 as a negative regulator of vacuolar fusion. Moreover, it suggests that this lipid acts as a molecular switch between fission and fusion.

Author response for "Copper blocks V‐ATPase activity and SNARE complex formation to inhibit yeast vacuole fusion"

2019 ◽  
Author(s):  
Gregory E. Miner ◽  
Katherine D. Sullivan ◽  
Chi Zhang ◽  
Logan R. Hurst ◽  
Matthew L. Starr ◽  
...  
Keyword(s):  
Complex Formation ◽  
Atpase Activity ◽  
Snare Complex ◽  
Author Response ◽  
Yeast Vacuole ◽  
Vacuole Fusion ◽  
Snare Complex Formation
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