Kinetics of endocytosis and recycling of the GPI-anchored variant surface glycoprotein in Trypanosoma brucei
Kinetics of endocytosis and recycling of the GPI-anchored variant surface glycoprotein in Trypanosoma brucei
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DOI：
10.1242/jcs.00938
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发表时间：
2004-03-01
影响因子：
4
通讯作者：
Overath, P
中科院分区：
文献类型：
--
作者：
Engstler, M;Thilo, L;Overath, P
覆盖寄生非洲锥虫的糖基磷脂酰肌醇 (GPI) 锚定变异表面糖蛋白 (VSG) 的致密外壳对于哺乳动物宿主的生存至关重要。 VSG 完全通过质膜的一个特殊部分（鞭毛袋）内化和回收。对 VSG 内吞作用和再循环动力学的直接测量表明，VSG 细胞表面池在 12 分钟内完成了翻转。相应地，细胞内池的周转（总 VSG 的 9 +/- 4%）仅需要 1 分钟，考虑到胞吞作用和胞吐作用仅限于细胞表面积的 5%，这是一个非常高的速率。使用生物素化 VSG 和一组区室标记进行的动力学 3D 共定位分析为 VSG 在细胞中的行程提供了一致的证据：VSG 在网格蛋白包被的大囊泡中被内吞，这些囊泡以每秒 6-7 个囊泡的速度从鞭毛袋膜中出芽，然后被递送到 RAB5 阳性早期内体。从那里，VSG 在两个阶段直接或通过 RAB7 阳性晚期内体再循环至 RAB11 阳性再循环内体。网格蛋白包被的小囊泡携带液相货物，并从早期和回收内体中耗尽 VSG 芽。假定这些囊泡将其内容物递送至晚期内体和/或溶酶体。回收内体产生 RAB11 阳性胞吐载体，与鞭毛袋融合，从而将 VSG 返回到细胞表面。 VSG 回收为 GPI 锚定蛋白的细胞运输和分类研究提供了一个有趣的模型。
The dense coat of glycosylphosphatidylinositol (GPI)anchored variant surface glycoprotein (VSG) covering parasitic African trypanosomes is essential for survival in mammalian hosts. VSG is internalised and recycled exclusively via a specialised part of the plasma membrane, the flagellar pocket. Direct measurement of the kinetics of VSG endocytosis and recycling shows that the VSG cell-surface pool is turned over within 12 minutes. Correspondingly, the turnover of the intracellular pool (9 +/- 4% of total VSG) requires only 1 minute, and this is an exceptionally high rate considering that endocytosis and exocytosis are limited to only 5% of the cell surface area. Kinetic 3D co-localisation analysis using biotinylated VSG and a panel of compartmental markers provides consistent evidence for the itinerary of VSG through the cell: VSG is endocytosed in large clathrin-coated vesicles, which bud from the flagellar pocket membrane at a rate of 6-7 vesicles per second, and is then delivered to RAB5-positive early endosomes. From there, VSG is recycled to RAB11-positive recycling endosomes at two stages, either directly or via RAB7-positive, late endosomes. Small clathrin-coated vesicles carrying fluid-phase cargo and being depleted of VSG bud from early and recycling endosomes. These vesicles are postulated to deliver their content to late endosomes and/or the lysosome. The recycling endosomes give rise to RAB11-positive exocytic carriers that fuse with the flagellar pocket and thereby return VSG to the cell surface. VSG recycling provides an interesting model for studies on the cellular trafficking and sorting of GPI-anchored proteins.