Sinai A P, Joiner K A
Infectious Diseases Section, Department of Internal Medicine, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA.
J Cell Biol. 2001 Jul 9;154(1):95-108. doi: 10.1083/jcb.200101073.
Toxoplasma gondii replicates within a specialized vacuole surrounded by the parasitophorous vacuole membrane (PVM). The PVM forms intimate interactions with host mitochondria and endoplasmic reticulum (ER) in a process termed PVM-organelle association. In this study we identify a likely mediator of this process, the parasite protein ROP2. ROP2, which is localized to the PVM, is secreted from anterior organelles termed rhoptries during parasite invasion into host cells. The NH(2)-terminal domain of ROP2 (ROP2hc) within the PVM is exposed to the host cell cytosol, and has characteristics of a mitochondrial targeting signal. In in vitro assays, ROP2hc is partially translocated into the mitochondrial outer membrane and behaves like an integral membrane protein. Although ROP2hc does not translocate across the ER membrane, it does exhibit carbonate-resistant binding to this organelle. In vivo, ROP2hc expressed as a soluble fragment in the cytosol of uninfected cells associates with both mitochondria and ER. The 30-amino acid (aa) NH(2)-terminal sequence of ROP2hc, when fused to green fluorescent protein (GFP), is sufficient for mitochondrial targeting. Deletion of the 30-aa NH(2)-terminal signal from ROP2hc results in robust localization of the truncated protein to the ER. These results demonstrate a new mechanism for tight association of different membrane-bound organelles within the cell cytoplasm.
刚地弓形虫在由寄生泡膜（PVM）包围的特殊泡内进行复制。在一个称为PVM-细胞器关联的过程中，PVM与宿主线粒体和内质网（ER）形成紧密的相互作用。在本研究中，我们鉴定出了这一过程的一个可能的介导因子，即寄生虫蛋白ROP2。ROP2定位于PVM，在寄生虫侵入宿主细胞期间从前称作棒状体的细胞器分泌。PVM内ROP2的NH(2)-末端结构域（ROP2hc）暴露于宿主细胞质溶胶中，并具有线粒体靶向信号的特征。在体外试验中，ROP2hc部分转运至线粒体外膜，其行为类似于整合膜蛋白。虽然ROP2hc不会穿过ER膜转运，但它确实表现出与该细胞器的抗碳酸盐结合。在体内，在未感染细胞的细胞质中作为可溶性片段表达的ROP2hc与线粒体和ER均有关联。当与绿色荧光蛋白（GFP）融合时，ROP2hc的30个氨基酸（aa）的NH(2)-末端序列足以实现线粒体靶向。从ROP2hc中删除30个氨基酸的NH(2)-末端信号会导致截短蛋白在ER上的稳定定位。这些结果证明了细胞质内不同膜结合细胞器紧密关联的一种新机制。
