The size and copy number of an organelle depend on an equilibrium of membrane fusion and fission. In vitro reconstitution of yeast vacuole fission and fusion shows that TORC1 selectively stimulates fission but does not change fusion activity. This explains the morphological transitions of yeast vacuoles in response to nutrient availability.

The equilibrium of membrane fusion and fission influences the volume and copy number of organelles. Fusion of yeast vacuoles has been well characterized but their fission and the mechanisms determining vacuole size and abundance remain poorly understood. We therefore attempted to systematically characterize factors necessary for vacuole fission. Here, we present results of an in vivo screening for deficiencies in vacuolar fragmentation activity of an ordered collection deletion mutants, representing 4881 non-essential genes of the yeast Saccharomyces cerevisiae. The screen identified 133 mutants with strong defects in vacuole fragmentation. These comprise numerous known fragmentation factors, such as the Fab1p complex, Tor1p, Sit4p and the V-ATPase, thus validating the approach. The screen identified many novel factors promoting vacuole fragmentation. Among those are 22 open reading frames of unknown function and three conspicuous clusters of proteins with known function. The clusters concern the ESCRT machinery, adaptins, and lipases, which influence the production of diacylglycerol and phosphatidic acid. A common feature of these factors of known function is their capacity to change membrane curvature, suggesting that they might promote vacuole fragmentation via this property.

Background: Contribution of SNARE transmembrane domains (TMDs) to membrane fusion is unclear. Results: Exchange of yeast vacuolar SNARE TMDs against unrelated TMDs yielded normal fusion activities; lipid anchors do not support fusion pore opening. Conclusion: SNARE TMDs serve as nonspecific membrane anchors in vacuole fusion but they need to be proteinaceous. Significance: SNARE TMDs participate in fusion pore opening.

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