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Vol. 13, Issue 3, 782-794, March 2002
Department of Biochemistry, Dartmouth Medical School, Hanover, New
Hampshire 03755-3844
Yeast vacuoles undergo fission and homotypic fusion, yielding one
to three vacuoles per cell at steady state. Defects in vacuole fusion
result in vacuole fragmentation. We have screened 4828 yeast strains,
each with a deletion of a nonessential gene, for vacuole morphology
defects. Fragmented vacuoles were found in strains deleted for genes
encoding known fusion catalysts as well as 19 enzymes of lipid
metabolism, 4 SNAREs, 12 GTPases and GTPase effectors, 9 additional known vacuole protein-sorting genes, 16 protein kinases, 2 phosphatases, 11 cytoskeletal proteins, and 28 genes of unknown
function. Vacuole fusion and vacuole protein sorting are catalyzed by
distinct, but overlapping, sets of proteins. Novel pathways of vacuole
priming and docking emerged from this deletion screen. These include
ergosterol biosynthesis, phosphatidylinositol (4,5)-bisphosphate turnover, and signaling from Rho GTPases to actin
remodeling. These pathways are supported by the sensitivity of the late
stages of vacuole fusion to inhibitors of phospholipase C, calcium
channels, and actin remodeling. Using databases of yeast protein
interactions, we found that many nonessential genes identified in our
deletion screen interact with essential genes that are directly
involved in vacuole fusion. Our screen reveals regulatory pathways of
vacuole docking and provides a genomic basis for studies of this reaction.
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