As a result of the genome sequencing project in Saccharomyces cerevisiae, three open reading frames were found in the yeast genome that contain sequences with strong homology to all the domains conserved among the four mammalian phosphatidylinositol-phosphate 5-phosphatases: inpp5bp, ocrl1p, synaptojanin, and ship. In addition, all three yeast gene products shared with synaptojanin regions of homology to the SAC1 gene of yeast. Disruption of each of these genes singly and in pairs produced mutant strains that were viable but demonstrated variable phenotypes of abnormal vacuolar and plasma membrane morphology as well as increased sensitivity to osmotic stress. Total phosphatidylinositol-(4,5)-bisphosphate 5-phosphatase activity was reduced to varying degrees in each of the strains. No defect in carboxypeptidase Y sorting was seen in a processing and targeting assay. Abnormal actin cytoskeleton morphology was present in some of the strains carrying mutations in two of the genes.