Toxoplasma gondii MAF1b Binds the Host Cell MIB Complex To Mediate Mitochondrial Association
Toxoplasma gondii MAF1b Binds the Host Cell MIB Complex To Mediate Mitochondrial Association
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DOI：
10.1128/msphere.00183-17
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发表时间：
2017-05-01
期刊：
影响因子：
4.8
通讯作者：
Boothroyd, John C.
中科院分区：
文献类型：
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作者：
Kelly, Felice D.;Wei, Brian M.;Boothroyd, John C.
许多不同的细胞内病原体，如嗜肺军团菌、鹦鹉热衣原体、脑胞内原虫，和弓形虫，操纵和重新定位宿主细胞器，包括线粒体。弓形虫速殖子使用分泌的蛋白质，线粒体结合因子1b（MAF 1b），驱动宿主线粒体和寄生虫空泡膜之间的结合，寄生虫在其中生长。然而，在这种相互作用中的宿主伙伴的身份以前没有被确定。通过在小鼠胚胎成纤维细胞中外源表达标记的MAF 1b，我们能够分离出与MAF 1b特异性相互作用的宿主细胞蛋白。然后，我们在MAF 1b表达成纤维细胞中以及在人成纤维细胞和HeLa细胞中寄生虫感染的背景下验证了这些相互作用。结果表明，宿主细胞线粒体复合物，线粒体膜间空间桥接（MIB）复合物，特异性地与MAF 1b相互作用。我们进一步证明，MAF 1b的一个版本，是缺乏在主机线粒体协会不能有效地共沉淀的MIB复合物。MAF 1b-MIB相互作用的重要性的验证来自于显示两个MIB复合物组分MIC 60和SAM 50的敲低，大大减少了线粒体与寄生虫空泡膜的结合。寄生虫分泌的膜整合蛋白和宿主细胞器的膜结合复合物之间的这种相互作用代表了宿主和病原体分子都已知的细胞器重新定位的第一个实例，并为更详细的生化研究提供了基础。常见的人类寄生虫弓形虫只在宿主细胞的空泡中复制，并通过分泌蛋白质改变其宿主细胞的环境。其中一种分泌蛋白MAF 1b的作用是将线粒体集中在寄生虫的空泡周围，这种重新定位改变了宿主的免疫反应。许多其他细胞内病原体也招募宿主线粒体，但介导这种相互作用的伴侣的身份以前没有在任何感染中描述过。在这里，我们表明，弓形虫MAF 1b结合到多功能的MIB蛋白复合物的主机线粒体。降低线粒体复合体中的蛋白质水平会降低宿主细胞线粒体和寄生虫液泡的紧密联系。这项工作为关键的宿主-病原体相互作用提供了新的见解，并确定了未来治疗干预的可能靶点，以及对重要生物学的更多分子理解。
Many diverse intracellular pathogens, such as Legionella pneumophila, Chlamydia psittaci, Encephalitozoon sp., and Toxoplasma gondii, manipulate and relocate host cell organelles, including mitochondria. Toxoplasma tachyzoites use a secreted protein, mitochondrial association factor 1b (MAF1b), to drive the association between the host mitochondria and the membrane of the parasitophorous vacuole, in which the parasites grow. The identity of the host partner in this interaction, however, has not previously been identified. By exogenously expressing tagged MAF1b in mouse embryonic fibroblasts, we were able to isolate host cell proteins that specifically interact with MAF1b. We then verified these interactions in the MAF1b-expressing fibroblasts, as well as in the context of parasite infection in human fibroblasts and HeLa cells. The results show that a host cell mitochondrial complex, the mitochondrial intermembrane space bridging (MIB) complex, specifically interacts with MAF1b. We further demonstrate that a version of MAF1b that is deficient in host-mitochondrial association does not efficiently coprecipitate the MIB complex. Validation of the importance of the MAF1b-MIB interaction came from showing that knockdown of two MIB complex components, MIC60 and SAM50, substantially reduces mitochondrial association with the parasitophorous vacuole membrane. This interaction between a secreted membrane-integral parasite protein and a membrane-bound complex of a host organelle represents the first instance of organelle relocalization in which both the host and pathogen molecules are known and provides the foundation for more detailed biochemical studies.IMPORTANCE Parasites interact intimately with their hosts, and the interactions shape both parties. The common human parasite Toxoplasma gondii replicates exclusively in a vacuole in a host cell and alters its host cell's environment through secreted proteins. One of these secreted proteins, MAF1b, acts to concentrate mitochondria around the parasite's vacuole, and this relocalization alters the host immune response. Many other intracellular pathogens also recruit host mitochondria, but the identities of the partners that mediate this interaction have not previously been described in any infection. Here, we show that Toxoplasma MAF1b binds to the multifunctional MIB protein complex on the host mitochondria. Reducing the levels of the proteins in this mitochondrial complex reduces the close association of host cell mitochondria and the parasite's vacuole. This work provides new insight into a key host-pathogen interaction and identifies possible targets for future therapeutic intervention as well as a more molecular understanding of important biology.