Hay J C, Fisette P L, Jenkins G H, Fukami K, Takenawa T, Anderson R A, Martin T F
Program in Cell and Molecular Biology, University of Wisconsin, Madison 53706.
Nature. 1995 Mar 9;374(6518):173-7. doi: 10.1038/374173a0.
Regulated fusion of secretory granules with the plasma membrane in secretory cells requires ATP, Ca2+ and cytosolic as well as membrane proteins. ATP-dependent steps in Ca(2+)-activated secretion from PC12 cells require three cytosolic PEP proteins (priming in exocytosis proteins, PEP1-3), the identity of which will provide insights into the required ATP-using reactions. PEP3 was recently identified as phosphatidylinositol transfer protein (PtdInsTP), and here we report that PEP1 consists of the type I phosphatidylinositol-4-phosphate 5-kinase (PtdInsP5K). The roles of PEP3/PtdInsTP and PEP1/PtdInsP5K in sequential phosphoinositide recruitment and phosphorylation explains their synergistic activity in ATP-dependent priming. Moreover, inhibition of Ca(2+)-activated secretion by PtdIns(4,5)P2-specific antibodies and phospholipase C implies that 5-phosphorylated inositides play a novel, necessary role in the regulated secretory pathway. The results indicate that lipid kinase-mediated phosphorylation is an important basis for ATP use in the exocytotic pathway.
分泌细胞中分泌颗粒与质膜的调控融合需要ATP、Ca2+、胞质蛋白以及膜蛋白。PC12细胞中Ca(2+)激活分泌过程中依赖ATP的步骤需要三种胞质PEP蛋白（胞吐作用起始蛋白，PEP1 - 3），其身份将为所需的ATP利用反应提供见解。PEP3最近被鉴定为磷脂酰肌醇转移蛋白（PtdInsTP），在此我们报告PEP1由I型磷脂酰肌醇 - 4 - 磷酸5 - 激酶（PtdInsP5K）组成。PEP3/PtdInsTP和PEP1/PtdInsP5K在磷酸肌醇的顺序募集和磷酸化中的作用解释了它们在ATP依赖起始过程中的协同活性。此外，PtdIns(4,5)P2特异性抗体和磷脂酶C对Ca(2+)激活分泌的抑制表明5 - 磷酸化肌醇在调控分泌途径中起新的必要作用。结果表明脂质激酶介导的磷酸化是胞吐途径中ATP利用的重要基础。
