Desrivières S, Cooke F T, Parker P J, Hall M N
Department of Biochemistry, Biozentrum, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
J Biol Chem. 1998 Jun 19;273(25):15787-93. doi: 10.1074/jbc.273.25.15787.
The Saccharomyces cerevisiae protein MSS4 is essential and homologous to mammalian phosphatidylinositol-4-phosphate (PI(4)P) 5-kinases. Here, we demonstrate that MSS4 is a lipid kinase. MSS4 has dual substrate specificity in vitro, converting PI(4)P to PI(4, 5)P2 and to a lesser extent PI(3)P to PI(3,4)P2; no activity was detected with PI or PI(5)P as a substrate. Cells overexpressing MSS4 contain an elevated level specifically of PI(4,5)P2, whereas mss4 mutant cells have only approximately 10% of the normal amount of this phosphorylated phosphoinositide. Furthermore, cells lacking MSS4 are unable to form actin cables and to properly localize their actin cytoskeleton during polarized cell growth. Overexpression of RHO2, encoding a Rho-type GTPase involved in regulation of the actin cytoskeleton, restores growth and polarized distribution of actin in an mss4 mutant. These results suggest that MSS4 is the major PI(4)P 5-kinase in yeast and provide a link between phosphoinositide metabolism and organization of the actin cytoskeleton in vivo.
酿酒酵母蛋白MSS4是必需的，且与哺乳动物磷脂酰肌醇-4-磷酸（PI(4)P）5-激酶同源。在此，我们证明MSS4是一种脂质激酶。MSS4在体外具有双重底物特异性，可将PI(4)P转化为PI(4,5)P2，在较小程度上也可将PI(3)P转化为PI(3,4)P2；以PI或PI(5)P作为底物时未检测到活性。过表达MSS4的细胞中PI(4,5)P2的水平显著升高，而mss4突变细胞中这种磷酸化磷脂酰肌醇的含量仅为正常水平的约10%。此外，缺乏MSS4的细胞在极化细胞生长过程中无法形成肌动蛋白丝束，也无法正确定位其肌动蛋白细胞骨架。编码参与肌动蛋白细胞骨架调节的Rho型GTP酶的RHO2的过表达可恢复mss4突变体中的生长以及肌动蛋白的极化分布。这些结果表明MSS4是酵母中的主要PI(4)P 5-激酶，并在体内磷酸肌醇代谢与肌动蛋白细胞骨架的组织之间建立了联系。
