Distinct roles for the yeast phosphatidylinositol 4-kinases, Stt4p and Pik1p, in secretion, cell growth, and organelle membrane dynamics
Distinct roles for the yeast phosphatidylinositol 4-kinases, Stt4p and Pik1p, in secretion, cell growth, and organelle membrane dynamics
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DOI：
10.1091/mbc.11.8.2673
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发表时间：
2000-08-01
影响因子：
3.3
通讯作者：
Emr, SD
中科院分区：
文献类型：
--
作者：
Audhya, A;Foti, M;Emr, SD
酵母酿酒酵母具有两个编码磷脂酰肌醇（PtdIns）4-激酶的基因STT 4和PIK 1。这两种基因产物在肌醇环的D-4位置磷酸化PtdIns以产生PtdIns（4）P，其在酵母活力中起重要作用，因为STT 4或PIK 1的缺失是致命的。此外，虽然这两种酶具有相同的生物化学活性，但任何一种激酶的表达增加都不能补偿另一种激酶的损失，这表明这些激酶调节不同的细胞内功能，每种功能都是酵母细胞生长所需的。通过构建温度条件下的单突变体和双突变体，我们发现Stt 4p活性是维持液泡形态、细胞壁完整性和肌动蛋白细胞骨架组织所必需的。相反，Pik 1 p是必不可少的正常分泌，高尔基体和液泡膜动力学，和内吞作用。引人注目的是，pik 1（ts）细胞在高尔基体修饰的分泌途径货物、Hsp 150 p和转化酶的分泌中表现出快速缺陷，而stt 4（ts）细胞表现出未检测到的分泌缺陷。两种单突变体都使PtdIns（4）P减少了约50%;然而，stt 4（ts）/pik 1（ts）双突变体细胞产生的PtdIns（4）P和PtdIns（4，5）P-2减少了10倍以上。在pik 1（ts）突变体中发现的异常高尔基体形态与缺乏Arf 1 p功能的细胞中发现的异常高尔基体形态惊人地相似，Arf 1 p是一种小的GTlp，已知其调节整个细胞的多个膜运输事件。与这一观察结果一致，arf 1突变体表现出：PtdIns（4）P水平降低。相比之下，在限制性温度下，在stt 4（ts）细胞中观察到的PtdIns（4）P水平下降，导致液泡大小与pik 1（ts）细胞相比发生显着变化，并导致肌动蛋白持续离域。基于这些结果，我们提出，Stt 4p和Pik 1 p作为主要的，如果不是唯一的，PtdIns 4-激酶在酵母中，并产生不同的池PtdIns（4）P和PtdIns（4，5）P-2，作用于不同的细胞内膜招募或激活尚未表征的效应蛋白。
The yeast Saccharomyces cerevisiae possesses two genes that encode phosphatidylinositol (PtdIns) 4-kinases, STT4 and PIK1. Both gene products phosphorylate PtdIns at the D-4 position of the inositol ring to generate PtdIns(4)P, which plays an essential role in yeast viability because deletion of either STT4 or PIK1 is lethal. Furthermore, although both enzymes have the same biochemical activity, increased expression of either kinase cannot compensate for the loss of the other, suggesting that these kinases regulate distinct intracellular functions, each of which is required for yeast cell growth. By the construction of temperature-conditional single and double mutants, we have found that Stt4p activity is required for the maintenance of vacuole morphology, cell wall integrity, and actin cytoskeleton organization. In contrast, Pik1p is essential for normal secretion, Golgi and vacuole membrane dynamics, and endocytosis. Strikingly, pik1(ts) cells exhibit a rapid defect in secretion of Golgi-modified secretory pathway cargos, Hsp150p and invertase, whereas stt4(ts) cells exhibit no detectable secretory defects. Both single mutants reduce PtdIns(4)P by similar to 50%; however, stt4(ts)/pik1(ts) double mutant cells produce more than 10-fold less PtdIns(4)P as well as PtdIns(4,5)P-2. The aberrant Golgi morphology found in pik1(ts) mutants is strikingly similar to that found in cells lacking the function of Arf1p, a small GTPase that is known to regulate multiple membrane trafficking events throughout the cell. Consistent with this observation, arf1 mutants exhibit: reduced PtdIns(4)P levels. In contrast, diminished levels of PtdIns(4)P observed in stt4(ts) cells at restrictive temperature result in a dramatic change in vacuole size compared with pik1(ts) cells and persistent actin delocalization. Based on these results, we propose that Stt4p and Pik1p act as the major, if not the only, PtdIns 4-kinases in yeast and produce distinct pools of PtdIns(4)P and PtdIns(4,5)P-2 that act on different intracellular membranes to recruit or activate as yet uncharacterized effector proteins.