SAC1 encodes a regulated lipid phosphoinositide phosphatase, defects in which can be suppressed by the homologous Inp52p and Inp53p phosphatases

The yeast protein Sac1p is involved in a range of cellular functions, inclu ding inositol metabolism, actin cytoskeletal organization, endoplasmic reti culum ATP transport, phosphatidylinositol-phosphatidylcholine transfer prot ein function, and multiple-drug sensitivity. The activity of Sac1p and its relationship to these phenotypes are unresolved. We show here that the regu lation of lipid phosphoinositides in sac1 mutants is defective, resulting i n altered levels of all lipid phosphoinositides, particularly phosphatidyli nositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate. We have iden tified two proteins with homology to Sac1p that can suppress drug sensitivi ty and also restore the levels of the phosphoinositides in sac1 mutants. Ov erexpression of truncated forms of these suppressor genes confirmed that su ppression was due to phosphoinositide phosphatase activity within these pro teins. We have now demonstrated this activity for Sac1p and have characteri zed its specificity. The in vitro phosphatase activity and specificity of S ac1p were not altered by some mutations. Indeed, in vivo mutant Sac1p phosp hatase activity also appeared unchanged under conditions in which cells wer e drug-resistant. However, under different growth conditions, both drug sen sitivity and the phosphatase defect were manifest. It is concluded that SAC 1 encodes a novel lipid phosphoinositide phosphatase in which specific muta tions can cause the sac1 phenotypes by altering the in vivo regulation of t he protein rather than by destroying phosphatase activity.