Kim S J, Rahbar R, Hegde R S
Laboratory of Cellular Oncology, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA.
J Biol Chem. 2001 Jul 13;276(28):26132-40. doi: 10.1074/jbc.M101638200. Epub 2001 May 18.
The prion protein (PrP) is synthesized in three topologic forms at the endoplasmic reticulum. (sec)PrP is fully translocated into the endoplasmic reticulum lumen, whereas (Ntm)PrP and (Ctm)PrP are single-spanning membrane proteins of opposite orientation. Increased generation of (Ctm)PrP in either transgenic mice or humans is associated with the development of neurodegenerative disease. To study the mechanisms by which PrP can achieve three topologic outcomes, we analyzed the translocation of proteins containing mutations introduced into either the N-terminal signal sequence or potential transmembrane domain (TMD) of PrP. Although mutations in either domain were found to affect PrP topogenesis, they did so in qualitatively different ways. In addition to its traditional role in mediating protein targeting, the signal was found to play a surprising role in determining orientation of the PrP N terminus. By contrast, the TMD was found to influence membrane integration. Analysis of various signal and TMD double mutants demonstrated that the topologic consequence of TMD action was directly dependent on the previous, signal-mediated step. Together, these results reveal that PrP topogenesis is controlled at two discrete steps during its translocation and provide a framework for understanding how these steps act coordinately to determine the final topology achieved by PrP.
朊病毒蛋白（PrP）在内质网以三种拓扑形式合成。（sec）PrP完全转运到内质网腔中，而（Ntm）PrP和（Ctm）PrP是方向相反的单次跨膜蛋白。在转基因小鼠或人类中，（Ctm）PrP生成增加与神经退行性疾病的发展相关。为了研究PrP实现三种拓扑结果的机制，我们分析了含有引入PrP N端信号序列或潜在跨膜结构域（TMD）突变的蛋白质的转运。尽管发现任一结构域中的突变都会影响PrP的拓扑发生，但它们的影响方式在质上有所不同。除了其在介导蛋白质靶向中的传统作用外，还发现该信号在确定PrP N端方向方面发挥了惊人的作用。相比之下，发现TMD影响膜整合。对各种信号和TMD双突变体的分析表明，TMD作用的拓扑结果直接取决于先前的信号介导步骤。总之，这些结果表明PrP的拓扑发生在其转运过程中的两个离散步骤受到控制，并为理解这些步骤如何协同作用以确定PrP最终实现的拓扑结构提供了一个框架。
