Chitama Ben-Yeddy Abel, Miyazaki Shinya, Zhu Xiaotong, Kagaya Wataru, Yahata Kazuhide, Kaneko Osamu
Leading Program, Graduate School of Biomedical Sciences, Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523, Japan; Department of Protozoology, Institute of Tropical Medicine (NEKKEN), Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523, Japan.
Department of Protozoology, Institute of Tropical Medicine (NEKKEN), Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523, Japan; Leiden Malaria Research Group, Parasitology, Leiden University Medical Center (LUMC), Leiden, The Netherlands.
Parasitol Int. 2019 Aug;71:186-193. doi: 10.1016/j.parint.2019.04.019. Epub 2019 Apr 24.
Plasmodium falciparum, an obligate intracellular protozoan parasite which causes the severe form of human malaria, exports numerous proteins to the infected red blood cell that are important for its survival and of severe pathological effect to its host. These proteins and their export mechanisms are candidates for drug and vaccine development, and among them is the Plasmodium SURFIN family of proteins. Previously we showed that the N-terminal region along with the sequence surrounding the transmembrane domain of SURFIN is essential for its export to Maurer's clefts in the red blood cell cytoplasm. We proposed that this region is recognized by a machinery responsible for protein translocation across the parasitophorous vacuole membrane surrounding the parasite. To understand the export mechanism further, we utilized a fluorescent protein-tagged mini-SURFIN consisting of the minimum essential components for export. Alanine scanning of all charged amino acids within the N-terminal region revealed that replacement of 3 glutamic acid and 2 lysine residues significantly impairs the export efficiency of this protein across the parasitophorous vacuole membrane. In addition, N-terminally Myc-tagged mini-SURFIN and mini-SURFIN with similar architectures were detected with anti-Myc antibody at Maurer's clefts, indicating that elements required for export to Maurer's clefts are conserved between SURFIN and SURFIN, and that N-terminal sequences of these SURFIN members are not cleaved during export. Our results implicate a conserved nature of SURFIN export to the red blood cell, particularly an important role of multiple glutamic acid and lysine residues in the SURFIN N-terminal region.
恶性疟原虫是一种专性细胞内原生动物寄生虫，可导致严重的人类疟疾，它会向受感染的红细胞输出大量蛋白质，这些蛋白质对其生存至关重要，且对宿主具有严重的病理影响。这些蛋白质及其输出机制是药物和疫苗开发的候选对象，其中包括疟原虫SURFIN蛋白家族。此前我们表明，SURFIN的N端区域以及跨膜结构域周围的序列对于其输出到红细胞细胞质中的毛氏小体至关重要。我们推测该区域被负责蛋白质跨寄生虫周围的寄生泡膜转运的机制所识别。为了进一步了解输出机制，我们利用了一种荧光蛋白标记的微型SURFIN，它由输出所需的最小基本成分组成。对N端区域内所有带电荷氨基酸进行丙氨酸扫描发现，3个谷氨酸和2个赖氨酸残基的替换显著损害了该蛋白跨寄生泡膜的输出效率。此外，在毛氏小体中用抗Myc抗体检测到了N端带有Myc标签的微型SURFIN和具有相似结构的微型SURFIN，这表明输出到毛氏小体所需的元件在SURFIN之间是保守的，并且这些SURFIN成员的N端序列在输出过程中不会被切割。我们的结果表明SURFIN输出到红细胞具有保守性，特别是SURFIN N端区域中多个谷氨酸和赖氨酸残基的重要作用。
