Novel targeting signals mediate the sorting of different isoforms of the tail-anchored membrane protein cytochrome b5 to either endoplasmic reticulum or mitochondria
Novel targeting signals mediate the sorting of different isoforms of the tail-anchored membrane protein cytochrome b5 to either endoplasmic reticulum or mitochondria
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DOI：
10.1105/tpc.104.026039
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发表时间：
2004-11-01
期刊：
影响因子：
11.6
通讯作者：
Mullen, RT
中科院分区：
文献类型：
--
作者：
Hwang, YT;Pelitire, SM;Mullen, RT
尾锚定膜蛋白是一类通过位于C末端附近的单个疏水氨基酸片段整合到各种细胞器上的蛋白质。虽然在植物细胞中的特定亚细胞区室中的尾锚定蛋白的定位对于它们的生物学功能是必不可少的，但负责分选这些蛋白的分子靶向信号并没有很好地定义。在这里，我们描述了四个密切相关的桐（油桐）细胞色素B（5）亚型（Cb 5-A，-B，-C和-D），这是一个小尾巴锚定蛋白，在许多细胞过程中发挥重要作用，包括脂质生物合成的生物发生。使用体内和体外试验的组合，我们表明，Cb 5-A，-B，和-C专门针对内质网（ER），而Cb 5-D是专门针对线粒体外膜。ER和线粒体Cb 5的综合突变分析显示，它们的C末端，包括跨膜结构域（TMD）和尾部区域，包含几个独特的物理化学和序列特异性特征，定义了细胞器特异性靶向基序。沿着TMD的沿着一面的亲水性氨基酸的组合、TMD的中间部分的分支含β-碳残基的富集和C-末端尾部的二元-R-R/K/H-x基序介导了Cb 5的线粒体靶向。相比之下，Cb 5的ER靶向主要取决于TMD的总长度和疏水性，尽管尾部的-R/H-x-Y/F-基序也是靶向决定簇。总的来说，所呈现的结果为尾锚定蛋白进入ER或线粒体靶向途径所需的早期生物遗传事件提供了重要的见解。
Tail-anchored membrane proteins are a class of proteins that are targeted posttranslationally to various organelles and integrated by a single segment of hydrophobic amino acids located near the C terminus. Although the localization of tail-anchored proteins in specific subcellular compartments in plant cells is essential for their biological function, the molecular targeting signals responsible for sorting these proteins are not well defined. Here, we describe the biogenesis of four closely related tung (Aleurites fordii) cytochrome b(5) isoforms (Cb5-A, -B, -C, and -D), which are small tail-anchored proteins that play an essential role in many cellular processes, including lipid biosynthesis. Using a combination of in vivo and in vitro assays, we show that Cb5-A, -B, and -C are targeted exclusively to the endoplasmic reticulum (ER), whereas Cb5-D is targeted specifically to mitochondrial outer membranes. Comprehensive mutational analyses of ER and mitochondrial Cb5s revealed that their C termini, including transmembrane domains (TMD) and tail regions, contained several unique physicochemical and sequence-specific characteristics that defined organelle-specific targeting motifs. Mitochondrial targeting of Cb5 was mediated by a combination of hydrophilic amino acids along one face of the TMD, an enrichment of branched beta-carbon-containing residues in the medial portion of the TMD, and a dibasic -R-R/K/H-x motif in the C-terminal tail. By contrast, ER targeting of Cb5 depended primarily upon the overall length and hydrophobicity of the TMD, although an -R/H-x-Y/F- motif in the tail was also a targeting determinant. Collectively, the results presented provide significant insight into the early biogenetic events required for entry of tail-anchored proteins into either the ER or mitochondrial targeting pathways.