Translocation of Dense Granule Effectors across the Parasitophorous Vacuole Membrane in Toxoplasma-Infected Cells Requires the Activity of ROP17, a Rhoptry Protein Kinase.
Translocation of Dense Granule Effectors across the Parasitophorous Vacuole Membrane in Toxoplasma-Infected Cells Requires the Activity of ROP17, a Rhoptry Protein Kinase.
复制标题
致密颗粒效应器在弓形虫感染细胞中穿过寄生液泡膜的易位需要 ROP17（一种棒状体蛋白激酶）的活性。
DOI：
10.1128/msphere.00276-19
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发表时间：
2019
期刊：
影响因子：
4.8
通讯作者：
Boothroyd,JohnC
中科院分区：
文献类型：
--
作者：
Panas,MichaelW;Ferrel,Abel;Naor,Adit;Tenborg,Elizabeth;Lorenzi,HernanA;Boothroyd,JohnC
弓形虫速殖子通过引入一大套颗粒和致密颗粒衍生的效应蛋白来发挥宿主细胞的功能。这些效应物通过不同的途径到达宿主细胞质：直接注射棒状病毒效应器和通过寄生质空泡膜(PVM)移位致密颗粒(GRA)效应器。最近已经部分阐明了这些GRA效应器的移位机制，揭示了三个组分，MYR1、MYR2和MYR3。为了确定是否可能涉及其他蛋白质，我们回到GRA易位缺陷突变体库，选择了一个部分缺陷的突变体库，这表明它可能位于编码该机制的一个新组件的基因中。令人惊讶的是，全基因组测序显示，编码一种已知的棒状蛋白的基因发生了错义突变，这种蛋白是一种名为ROP17的丝氨酸/苏氨酸蛋白激酶。ROP17位于感染细胞中PVM的宿主胞浆一侧，此前已知其具有磷酸化活性，从而使宿主免疫相关的GTP酶失活。在这里，我们证明了ROP17的零突变体或催化死亡突变体在跨PVM的GRA易位方面存在缺陷，但这种易位可以被感染同一宿主细胞的其他速殖子传递的ROP17拯救。这有力地证明了ROP17的S在调节GRA易位中的作用是在PVM的寄主胞液侧进行的，而不是在寄生虫或寄生性液泡的管腔内。这代表了一种全新的方式，在这种方式下，弓形虫的不同分泌部分协同调节宿主-寄生虫的相互作用。当弓形虫感染细胞时，它在细胞周围建立一个保护性的寄生液泡。虽然这种液泡提供了保护，但它也是影响和控制宿主细胞的寄生虫效应蛋白出口的障碍。我们在这里的发现，寄生虫棒状蛋白ROP17是这些效应蛋白输出所必需的，除了它在保护液泡方面的已知作用外，还为ROP17提供了一个独特的、新的功能。这将使我们能够进一步研究如何防止效应蛋白的出口，从而防止弓形虫有效地感染其动物和人类宿主。
Toxoplasma gondii tachyzoites co-opt host cell functions through introduction of a large set of rhoptry- and dense granule-derived effector proteins. These effectors reach the host cytosol through different means: direct injection for rhoptry effectors and translocation across the parasitophorous vacuolar membrane (PVM) for dense granule (GRA) effectors. The machinery that translocates these GRA effectors has recently been partially elucidated, revealing three components, MYR1, MYR2, and MYR3. To determine whether other proteins might be involved, we returned to a library of mutants defective in GRA translocation and selected one with a partial defect, suggesting it might be in a gene encoding a new component of the machinery. Surprisingly, whole-genome sequencing revealed a missense mutation in a gene encoding a known rhoptry protein, a serine/threonine protein kinase known as ROP17. ROP17 resides on the host cytosol side of the PVM in infected cells and has previously been known for its activity in phosphorylating and thereby inactivating host immunity-related GTPases. Here, we show that null or catalytically dead mutants of ROP17 are defective in GRA translocation across the PVM but that translocation can be rescued “intrans” by ROP17 delivered by other tachyzoites infecting the same host cell. This strongly argues that ROP17’s role in regulating GRA translocation is carried out on the host cytosolic side of the PVM, not within the parasites or lumen of the parasitophorous vacuole. This represents an entirely new way in which the different secretory compartments ofToxoplasmatachyzoites collaborate to modulate the host-parasite interaction.IMPORTANCEWhenToxoplasmainfects a cell, it establishes a protective parasitophorous vacuole surrounding it. While this vacuole provides protection, it also serves as a barrier to the export of parasite effector proteins that impact and take control of the host cell. Our discovery here that the parasite rhoptry protein ROP17 is necessary for export of these effector proteins provides a distinct, novel function for ROP17 apart from its known role in protecting the vacuole. This will enable future research into ways in which we can prevent the export of effector proteins, thereby preventingToxoplasmafrom productively infecting its animal and human hosts.