Dissection of Minimal Sequence Requirements for Rhoptry Membrane Targeting in the Malaria Parasite
Dissection of Minimal Sequence Requirements for Rhoptry Membrane Targeting in the Malaria Parasite
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DOI：
10.1111/j.1600-0854.2012.01394.x
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发表时间：
2012-10-01
期刊：
影响因子：
4.5
通讯作者：
Gilberger, Tim W.
中科院分区：
文献类型：
--
作者：
Cabrera, Ana;Herrmann, Susann;Gilberger, Tim W.
棒状体是属于顶端复合体进化枝的单细胞生物特有的特殊分泌细胞器。这些细胞器通过积累并随后分泌未知数量的介导宿主细胞进入的蛋白质，在宿主细胞的入侵过程中发挥关键作用。尽管它们发挥着重要作用，但人们对其生物发生、成分和靶向决定因素知之甚少。在这里，我们报告了一种保守的顶端复合体蛋白，称为犰狳重复（ARO）蛋白，我们将其定位于恶性疟原虫和弓形虫棒状体的胞质表面。我们证明前 20 个 N 端氨基酸足以用于菱形膜靶向。该蛋白质依赖肉豆蔻酰化和棕榈酰化基序进行膜附着。尽管这些脂质修饰是必要的，但它们不足以将 ARO 引导至视杆细胞膜。突变分析揭示了 ARO 前 20 个氨基酸中对棒状体膜附着起重要作用的其他残基：带正电荷的残基 R9 和 K14。有趣的是，R9 与负电荷的交换完全消除了膜附着，而 K14（以及较小程度的 K16）的交换仅改变其膜特异性。此外，在疟疾寄生虫的基因组中鉴定出 17 个蛋白质，预计其前 20 个 N 末端氨基酸会被肉豆蔻酰化和棕榈酰化。虽然大多数相应的 GFP 融合蛋白被运输到寄生虫质膜，但有两个被分选到顶端细胞器。有趣的是，这些蛋白质具有与 ARO 相似的基序。
Rhoptries are specialized secretory organelles characteristic of single cell organisms belonging to the clade Apicomplexa. These organelles play a key role in the invasion process of host cells by accumulating and subsequently secreting an unknown number of proteins mediating host cell entry. Despite their essential role, little is known about their biogenesis, components and targeting determinants. Here, we report on a conserved apicomplexan protein termed Armadillo Repeats-Only (ARO) protein that we localized to the cytosolic face of Plasmodium falciparum and Toxoplasma gondii rhoptries. We show that the first 20 N-terminal amino acids are sufficient for rhoptry membrane targeting. This protein relies on both myristoylation and palmitoylation motifs for membrane attachment. Although these lipid modifications are essential, they are not sufficient to direct ARO to the rhoptry membranes. Mutational analysis revealed additional residues within the first 20 amino acids of ARO that play an important role for rhoptry membrane attachment: the positively charged residues R9 and K14. Interestingly, the exchange of R9 with a negative charge entirely abolishes membrane attachment, whereas the exchange of K14 (and to a lesser extent K16) alters only its membrane specificity. Additionally, 17 proteins predicted to be myristoylated and palmitoylated in the first 20 N-terminal amino acids were identified in the genome of the malaria parasite. While most of the corresponding GFP fusion proteins were trafficked to the parasite plasma membrane, two were sorted to the apical organelles. Interestingly, these proteins have a similar motif identified for ARO.