Cell-cycle and developmental regulation of TbRAB31 localisation, a GTP-locked Rab protein from Trypanosoma brucei
Cell-cycle and developmental regulation of TbRAB31 localisation, a GTP-locked Rab protein from Trypanosoma brucei
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DOI：
10.1016/s0166-6851(99)00192-9
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发表时间：
2000-02-25
影响因子：
1.5
通讯作者：
Field, MC
中科院分区：
文献类型：
--
作者：
Field, H;Sherwin, T;Field, MC
Rab 蛋白是小型 GTP 酶，在膜区室之间的细胞内运输过程中控制囊泡融合的方向和时间。布氏锥虫的基因组测序和 EST 分析表明，锥虫 Rab (TbRAB) 基因家族介于酿酒酵母和哺乳动物之间，因此其细胞内转运途径的复杂性介于酿酒酵母和哺乳动物之间。 TbRAB31 是一种组成型表达的布氏锥虫 Rab 蛋白（以前称为 Trab7p），是两个紧密连锁的 TbRAB 基因之一的产物，另一个是 TbRAB2（TbRab2p，见：Field H、Ali BRS、Sherwin T、Gull K：Croft SL、Field MC。TbRab2p，布氏锥虫内质网的标记，定位于哺乳动物细胞中的ERGIC。J Cell Sci 1999.112.147-156）。参与 ER 到高尔基体的运输。 TbRAB31 与来自酿酒酵母的 Rab 蛋白 Sec4/Ypt1 亚家族的成员以及来自高等真核生物的 Rab13 和 Rab11 具有高度同源性。重组 TbRAB31 可结合 GTP，但与 Rab 蛋白不同的是，它具有不可检测的 GTP 酶活性，从而产生组成型 GTP 结合蛋白。 TbRAB31 抗体可识别位于间期原循环细胞动质体和细胞核之间的离散结构：相比之下，血流型 (BSF) 寄生虫的结构在形态上更为复杂，由至少两个病灶组成。 TbRAB31 在细胞周期中的行为也进行了研究； TbRAB31 总是定位于一个离散的结构，该结构在有丝分裂的早期复制并以与基体和动质体协调的方式重新定位到子细胞，这表明微管的参与。其他证据表明 TbRAB31 定位于锥虫高尔基复合体。首先，TbRAB31的间期位置与高尔基体位置一致。其次，TbRAB31 结构也可以被哺乳动物 β-外套蛋白 (β-COP) 的交叉反应抗体识别，该蛋白定位于哺乳动物细胞的高尔基体。第三，荧光神经酰胺类似物 BODIPY-TR-神经酰胺是哺乳动物高尔基体的可靠标记，与 TbRAB31 表现出重叠分布。通过使用二氨基联苯胺的组织化学和电子显微镜证实了 BODIPY-TR-神经酰胺在锥虫高尔基体中的位置。 (C) 2000 Elsevier Science B.V. 保留所有权利。
Rab proteins are small GTPases that control the direction and timing of vesicle fusion during intracellular trafficking between membraneous compartments. Genome sequencing and EST analysis of Trypanosoma brucei indicates that the trypanosome Rab (TbRAB) gene family, and hence complexity of intracellular transport pathways, is intermediate between Saccharomyces cerevisae and mammals. TbRAB31 is a constitutively expressed T. brucei Rab protein (formerly Trab7p) and is the product of one of two closely linked TbRAB genes, the other being TbRAB2 (TbRab2p, in: Field H, Ali BRS, Sherwin T, Gull K: Croft SL, Field MC. TbRab2p, a marker for the endoplasmic reticulum of Trypanosoma brucei, localises to the ERGIC in mammalian cells. J Cell Sci 1999.112.147-156). involved in ER to Golgi transport. TbRAB31 has high homology to members of the Sec4/Ypt1 subfamily of Rab proteins from S. cerevisiae and to Rab13 and Rab11 from higher eukaryotes. Recombinant TbRAB31 binds GTP but, unusually for a Rab protein, has undetectable GTPase activity resulting in a constitutively GTP-bound protein. Antibodies against TbRAB31 recognise a discrete structure located between the kinetoplast and nucleus in interphase procyclic cells: by contrast the structure is morphologically more complex in bloodstream form (BSF) parasites, consisting of at least two foci. TbRAB31 behaviour was also studied during the cell cycle; TbRAB31 always localised to a discrete structure that duplicated very early in mitosis and relocated to daughter cells in a coordinate manner with the basal body and kinetoplast, suggesting the involvement of microtubules. Additional evidence suggests that TbRAB31 localises to the trypanosome Golgi complex. Firstly, the interphase position of TbRAB31 is consistent with a Golgi location. Secondly, the TbRAB31 structure is also recognised by cross-reacting antibodies to mammalian beta-coatomer protein (beta-COP), which localises to the Golgi in mammalian cells. Thirdly, the fluorescent ceramide analogue, BODIPY-TR-ceramide, a reliable marker of the mammalian Golgi apparatus, exhibited overlapping distribution with TbRAB31. The location of BODIPY-TR-ceramide was confirmed at the trypanosome Golgi by histochemistry with diaminobenzidine and electron microscopy. (C) 2000 Elsevier Science B.V. All rights reserved.