Murakami Yoshiko, Siripanyapinyo Uamporn, Hong Yeongjin, Kang Ji Young, Ishihara Sonoko, Nakakuma Hideki, Maeda Yusuke, Kinoshita Taroh
Department of Immunoregulation, Research Institute for Microbial Diseases, Osaka University, Osaka 565-0871, Japan.
Mol Biol Cell. 2003 Oct;14(10):4285-95. doi: 10.1091/mbc.e03-03-0193. Epub 2003 Jun 13.
Many cell surface proteins are anchored to a membrane via a glycosylphosphatidylinositol (GPI), which is attached to the C termini in the endoplasmic reticulum. The inositol ring of phosphatidylinositol is acylated during biosynthesis of GPI. In mammalian cells, the acyl chain is added to glucosaminyl phosphatidylinositol at the third step in the GPI biosynthetic pathway and then is usually removed soon after the attachment of GPIs to proteins. The mechanisms and roles of the inositol acylation and deacylation have not been well clarified. Herein, we report derivation of human and Chinese hamster mutant cells defective in inositol acylation and the gene responsible, PIG-W. The surface expressions of GPI-anchored proteins on these mutant cells were greatly diminished, indicating the critical role of inositol acylation. PIG-W encodes a 504-amino acid protein expressed in the endoplasmic reticulum. PIG-W is most likely inositol acyltransferase itself because the tagged PIG-W affinity purified from transfected human cells had inositol acyltransferase activity and because both mutant cells were complemented with PIG-W homologs of Saccharomyces cerevisiae and Schizosaccharomyces pombe. The inositol acylation is not essential for the subsequent mannosylation, indicating that glucosaminyl phosphatidylinositol can flip from the cytoplasmic side to the luminal side of the endoplasmic reticulum.
许多细胞表面蛋白通过糖基磷脂酰肌醇（GPI）锚定在膜上，GPI在内质网中连接到蛋白的C末端。磷脂酰肌醇的肌醇环在GPI生物合成过程中被酰化。在哺乳动物细胞中，酰基链在GPI生物合成途径的第三步添加到氨基葡萄糖基磷脂酰肌醇上，然后通常在GPI与蛋白连接后不久被去除。肌醇酰化和去酰化的机制及作用尚未得到充分阐明。在此，我们报道了人及中国仓鼠肌醇酰化缺陷突变细胞的衍生以及相关基因PIG-W。这些突变细胞上GPI锚定蛋白的表面表达大幅减少，表明肌醇酰化的关键作用。PIG-W编码一种在内质网中表达的504个氨基酸的蛋白。PIG-W很可能本身就是肌醇酰基转移酶，因为从转染的人细胞中亲和纯化的带标签的PIG-W具有肌醇酰基转移酶活性，并且因为这两种突变细胞都能被酿酒酵母和粟酒裂殖酵母的PIG-W同源物互补。肌醇酰化对于随后的甘露糖基化并非必需，这表明氨基葡萄糖基磷脂酰肌醇可以从内质网的细胞质侧翻转到腔侧。
