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A more recent version of this article appeared on March 1, 2002
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Submitted on October 19, 2001
Revised on December 4, 2001
Accepted on December 18, 2001
1 Department of Biochemistry, Dartmouth Medical School, 7200 Vail Building, Hanover, NH 03755-3844
* Corresponding author. E-mail address: Bill.Wickner{at}Dartmouth.edu.
Yeast vacuoles undergo fission and homotypic fusion, yielding 1-3 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 (vam) 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 (VPS) genes, 16 protein kinases and 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 find that many non-essential genes identified in our deletion screen interact with essential genes which 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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