Sac1 lipid phosphatase and Stt4 phosphatidylinositol 4-kinase regulate a pool of phosphatidylinositol 4-phosphate that functions in the control of the actin cytoskeleton and vacuole morphology
Sac1 lipid phosphatase and Stt4 phosphatidylinositol 4-kinase regulate a pool of phosphatidylinositol 4-phosphate that functions in the control of the actin cytoskeleton and vacuole morphology
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DOI：
10.1091/mbc.12.8.2396
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发表时间：
2001-08-01
影响因子：
3.3
通讯作者：
Emr, SD
中科院分区：
文献类型：
--
作者：
Foti, M;Audhya, A;Emr, SD
磷酸肌醇的合成和周转是由一组最近鉴定的激酶和磷酸酶介导的严格调节的过程。我们分析了磷酸肌醇磷酸酶Sac 1 p在酿酒酵母中的主要作用，使用该基因的温度敏感等位基因。我们的分析表明，Sac 1 p的失活导致磷脂酰肌醇4-磷酸（PtdIns（4）P）的细胞水平的特定增加，伴随着液泡形态的变化和脂滴的积累。我们发现大部分Sac 1 p定位于内质网，这种定位对于PtdIns（4）P的有效周转至关重要。通过产生携带sac 1（ts）等位基因和两个温度敏感PtdIns 4-激酶基因stt 4（ts）或pik 1（ts）之一的双突变株，我们已经证明，在sac 1突变细胞中积累的大部分PtdIns（4）P是由Stt 4 PtdIns 4-激酶产生的，而不是Pik 1 p。与这些发现相一致，Sac 1 p的失活部分挽救了与stt 4 ts相关的缺陷，而不是pik 1（ts）突变细胞。为了分析Sac 1 p和其他同源磷酸肌醇磷酸酶sac 1（ts）之间的潜在重叠功能，产生了缺乏各种其他突触Janin样磷酸酶的突变体细胞。这些双重和三重突变体加剧了细胞内磷酸肌醇的积累，并导致高尔基体功能的缺陷。总之，我们的结果表明Saclp主要转化Stt 4p产生的PtdIns（4）P，并且Saclp的膜定位对其体内功能很重要。PtdIns（4）P库的调节对于液泡形态的维持、脂质储存的调节、高尔基体功能和肌动蛋白细胞骨架的组织似乎是至关重要的。
Synthesis and turnover of phosphoinositides are tightly regulated processes mediated by a set of recently identified kinases and phosphatases. We analyzed the primary role of the phosphoinositide phosphatase Sac1p in Saccharomyces cerevisiae with the use of a temperature-sensitive allele of this gene. Our analysis demonstrates that inactivation of Sac1p leads to a specific increase in the cellular levels of phosphatidylinositol 4-phosphate (PtdIns(4)P), accompanied by changes in vacuole morphology and an accumulation of lipid droplets. We have found that the majority of Sac1p localizes to the endoplasmic reticulum, and this localization is crucial for the efficient turnover of PtdIns(4)P. By generating double mutant strains harboring the sac1(ts) allele and one of two temperature-sensitive PtdIns 4-kinase genes, stt4(ts) or pik1(ts), we have demonstrated that the bulk of PtdIns(4)P that accumulates in sac1 mutant cells is generated by the Stt4 PtdIns 4-kinase, and not Pik1p. Consistent with these findings, inactivation of Sac1p partially rescued defects associated with stt4ts but not pik1(ts) mutant cells. To analyze potential overlapping functions between Sac1p and other homologous phosphoinositide phosphatases, sac1(ts), mutant cells lacking various other synaptojanin-like phosphatases were generated. These double and triple mutants exacerbated the accumulation of intracellular phosphoinositides and caused defects in Golgi function. Together, our results demonstrate that Saclp primarily turns over Stt4p-generated PtdIns(4)P and that the membrane localization of Saclp is important for its function in vivo. Regulation of this PtdIns(4)P pool appears to be crucial for the maintenance of vacuole morphology, regulation of lipid storage, Golgi function, and actin cytoskeleton organization.