Hutzler Johannes, Schmid Maria, Bernard Thomas, Henrissat Bernard, Strahl Sabine
Department of Cell Chemistry, Institute of Plant Sciences, University of Heidelberg, 69120 Heidelberg, Germany.
Proc Natl Acad Sci U S A. 2007 May 8;104(19):7827-32. doi: 10.1073/pnas.0700374104. Epub 2007 Apr 30.
Protein O-mannosylation represents an evolutionarily conserved, essential posttranslational modification with immense impact on a variety of cellular processes. In humans, O-mannosylation defects result in Walker-Warburg syndrome, a severe recessive congenital muscular dystrophy associated with defects in neuronal migration that produce complex brain and eye abnormalities. In mouse and yeasts, loss of O-mannosylation causes lethality. Protein O-mannosyltransferases (PMTs) initiate the assembly of O-mannosyl glycans. The evolutionarily conserved PMT family is classified into PMT1, PMT2, and PMT4 subfamilies, which mannosylate distinct target proteins. In contrast to other types of glycosylation, signal sequences for O-mannosylation have not been identified to date. In the present study, we identified signals that determine PMT4-dependent O-mannosylation. Using specific model proteins, we demonstrate that in yeast Pmt4p mediates O-mannosylation of Ser/Thr-rich membrane-attached proteins. The nature of the membrane-anchoring sequence is nonrelevant, as long as it is flanked by a Ser/Thr-rich domain facing the endoplasmic reticulum lumen. Our work shows that, in contrast to several other types of glycosylation, PMT4 O-mannosylation signals are not just linear protein's primary structure sequences but rather are highly complex. Based on these findings, we performed in silico analyses of the Saccharomyces cerevisiae proteome and identified previously undescribed Pmt4p substrates. This tool for proteome-wide identification of O-mannosylated proteins is of general interest because several of these proteins are major players of a wide variety of cellular processes.
蛋白质O-甘露糖基化是一种进化上保守的重要翻译后修饰，对多种细胞过程有巨大影响。在人类中，O-甘露糖基化缺陷会导致沃克-沃尔堡综合征，这是一种严重的隐性先天性肌营养不良症，与神经元迁移缺陷相关，会导致复杂的脑和眼异常。在小鼠和酵母中，O-甘露糖基化的缺失会导致致死性。蛋白质O-甘露糖基转移酶（PMT）启动O-甘露糖聚糖的组装。进化上保守的PMT家族分为PMT1、PMT2和PMT4亚家族，它们对不同的靶蛋白进行甘露糖基化。与其他类型的糖基化不同，迄今为止尚未鉴定出O-甘露糖基化的信号序列。在本研究中，我们鉴定了决定PMT4依赖性O-甘露糖基化的信号。使用特定的模型蛋白，我们证明在酵母中Pmt4p介导富含丝氨酸/苏氨酸的膜附着蛋白的O-甘露糖基化。膜锚定序列的性质无关紧要，只要它两侧是面向内质网腔的富含丝氨酸/苏氨酸的结构域。我们的工作表明，与其他几种类型的糖基化不同，PMT4 O-甘露糖基化信号不仅仅是线性蛋白质的一级结构序列，而是高度复杂的。基于这些发现，我们对酿酒酵母蛋白质组进行了计算机分析，并鉴定出先前未描述的Pmt4p底物。这种用于全蛋白质组鉴定O-甘露糖基化蛋白的工具具有普遍意义，因为这些蛋白中的几种是多种细胞过程的主要参与者。
