Abstract
In eukaryotic endomembrane systems, Qabc-SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) on one membrane and R-SNARE on the opposing membrane assemble into a trans-QabcR-SNARE complex to drive membrane fusion. However, it remains ambiguous whether pairing of Qabc- and R-SNAREs mediates membrane fusion specificity. Here, we explored the fusion specificity of reconstituted proteoliposomes bearing purified SNAREs in yeast vacuoles and other organelles. We found that not only vacuolar R-SNARE Nyv1p but also the non-cognate R-SNAREs, endosomal Snc2p, and endoplasmic reticulum-Golgi Sec22p caused efficient fusion with vacuolar Qabc-SNAREs. In contrast, their fusion is blocked completely by replacing vacuolar Qc-SNARE Vam7p with the non-cognate endosomal Tlg1p and Syn8p, although these endosomal Qc-SNAREs fully retained the ability to form cis-SNARE complexes with vacuolar SNAREs in solution and on membranes. Thus, our current study establishes that an appropriate assembly of Qabc-SNAREs is crucial for regulating fusion specificity, whereas R-SNARE itself has little contribution to specificity.
Highlights
Qabc- and R-sensitive factor attachment protein receptors (SNAREs) proteins are key components for membrane fusion in eukaryotic organelles
These studies show that vacuolar Qabc- and R-SNAREs undergo fusion when separately reconstituted into two reconstituted proteoliposomes (RPLs) populations [11, 12, 22, 28]
The current Qabc- and R-SNARE RPL preparations contained some amounts of a cleaved-off GST-His6 fragment (Fig. 1B), we have confirmed that a GST-His6 protein has little effect on our lipid mixing assays
Summary
Qabc- and R-SNARE proteins are key components for membrane fusion in eukaryotic organelles. Qabc-SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) on one membrane and R-SNARE on the opposing membrane assemble into a trans-QabcR-SNARE complex to drive membrane fusion. It remains ambiguous whether pairing of Qabc- and R-SNAREs mediates membrane fusion specificity. In vitro studies using reconstituted proteoliposomes (RPLs) indicate the intrinsic fusogenic capacities of isolated SNAREs, showing that pairs of Q-SNARE RPLs and R-SNARE RPLs can cause fusion through trans-SNARE complexes (19 –21) This RPL approach has been further employed to test whether SNAREs themselves ensure the specificity of membrane fusion [22,23,24]. We thoroughly compared the “cognate” set of Q- and R-SNAREs that is required for homotypic vacuole fusion with a variety of the “non-cognate” SNARE combinations and reveal the clear requirement for specific Qabc-SNARE assemblies for fusion
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
