ATG8 Is Essential Specifically for an Autophagy-Independent Function in Apicoplast Biogenesis in Blood-Stage Malaria Parasites.
ATG8 Is Essential Specifically for an Autophagy-Independent Function in Apicoplast Biogenesis in Blood-Stage Malaria Parasites.
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作者：
Walczak M;Ganesan SM;Niles JC;Yeh E
疟原虫寄生虫和相关病原体含有一种重要的非光合作用质体细胞器，即顶质体，源自次级内共生。有趣的是，一种高度保守的真核蛋白，自噬相关蛋白 8 (ATG8)，在顶质体中具有不依赖于自噬的功能。关于 ATG8 的新顶质体功能及其在血液阶段恶性疟原虫中的重要性，人们知之甚少。使用 ATG8 表达受到条件调节的恶性疟原虫菌株，我们发现恶性疟原虫 ATG8 (PfATG8) 对于寄生虫复制至关重要。值得注意的是，添加异戊烯焦磷酸 (IPP) 可以逆转由 PfATG8 缺失引起的生长抑制，此前研究表明异戊烯焦磷酸可以挽救恶性疟原虫的顶质体缺陷。缺乏PfATG8但其生长被IPP挽救的寄生虫已经失去了顶质体。我们设计了一套功能测定法，包括用于检测低拷贝数顶端质体基因组的新荧光原位杂交（FISH）方法，以询问顶端质体生物发生中的特定步骤并检测寄生虫复制受阻之前的顶端质体缺陷。尽管顶质体的蛋白质输入和膜扩张不受影响，但顶质体不会被子代寄生虫遗传。我们的研究结果表明，尽管 PfATG8 被提出具有多种自噬依赖性和独立功能，但只有其在顶端质体生物发生中的作用对于血液阶段寄生虫来说是必需的。我们认为 PfATG8 是寄生虫复制过程中顶质体裂变或分离所必需的。引起疟疾的疟原虫寄生虫和相关的顶复门寄生虫是重要的人类和兽医病原体。它们在进化上与传统模式生物相距甚远，并拥有独特的质体细胞器，即顶端质体，这是通过不寻常的真核生物-真核生物内共生获得的，这种内共生在寄生虫细胞中建立了新的蛋白质/脂质输入和细胞器遗传途径。尽管顶质体对于寄生虫生命周期各个阶段的生存至关重要，但人们对这些新的生物发生途径知之甚少。我们表明，疟疾寄生虫已经适应了酵母和哺乳动物中巨自噬所需的高度保守的蛋白质，以在顶质体遗传中特异性发挥作用。我们的发现阐明了致病真核生物这一不同分支中细胞器生物发生的新机制，这对于发病机制至关重要。
Plasmodium parasites and related pathogens contain an essential nonphotosynthetic plastid organelle, the apicoplast, derived from secondary endosymbiosis. Intriguingly, a highly conserved eukaryotic protein, autophagy-related protein 8 (ATG8), has an autophagy-independent function in the apicoplast. Little is known about the novel apicoplast function of ATG8 and its importance in blood-stage Plasmodium falciparum. Using a P. falciparum strain in which ATG8 expression was conditionally regulated, we showed that P. falciparum ATG8 (PfATG8) is essential for parasite replication. Significantly, growth inhibition caused by the loss of PfATG8 was reversed by addition of isopentenyl pyrophosphate (IPP), which was previously shown to rescue apicoplast defects in P. falciparum. Parasites deficient in PfATG8, but whose growth was rescued by IPP, had lost their apicoplast. We designed a suite of functional assays, including a new fluorescence in situ hybridization (FISH) method for detection of the low-copy-number apicoplast genome, to interrogate specific steps in apicoplast biogenesis and detect apicoplast defects which preceded the block in parasite replication. Though protein import and membrane expansion of the apicoplast were unaffected, the apicoplast was not inherited by daughter parasites. Our findings demonstrate that, though multiple autophagy-dependent and independent functions have been proposed for PfATG8, only its role in apicoplast biogenesis is essential in blood-stage parasites. We propose that PfATG8 is required for fission or segregation of the apicoplast during parasite replication. Plasmodium parasites, which cause malaria, and related apicomplexan parasites are important human and veterinary pathogens. They are evolutionarily distant from traditional model organisms and possess a unique plastid organelle, the apicoplast, acquired by an unusual eukaryote-eukaryote endosymbiosis which established novel protein/lipid import and organelle inheritance pathways in the parasite cell. Though the apicoplast is essential for parasite survival in all stages of its life cycle, little is known about these novel biogenesis pathways. We show that malaria parasites have adapted a highly conserved protein required for macroautophagy in yeast and mammals to function specifically in apicoplast inheritance. Our finding elucidates a novel mechanism of organelle biogenesis, essential for pathogenesis, in this divergent branch of pathogenic eukaryotes.