Rice SCAMP1 defines clathrin-coated, trans-Golgi-located tubular-vesicular structures as an early endosome in tobacco BY-2 cells
Rice SCAMP1 defines clathrin-coated, trans-Golgi-located tubular-vesicular structures as an early endosome in tobacco BY-2 cells
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DOI：
10.1105/tpc.106.045708
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发表时间：
2007-01-01
期刊：
影响因子：
11.6
通讯作者：
Jiang, Liwen
中科院分区：
文献类型：
--
作者：
Lam, Sheung Kwan;Siu, Ching Lung;Jiang, Liwen
我们最近发现多泡体（MVB）作为前液泡室（PVC）的分泌和内吞途径的裂解液泡在烟草（烟草）BY-2细胞。分泌载体膜蛋白（SCAMPs）是高尔基体后的膜蛋白，在动物细胞中介导内吞作用。为了确定植物中的内吞途径，我们克隆了动物SCAMP 1的水稻（Oryza sativa）同源物，并产生表达黄色荧光蛋白（YFP）-SCAMP 1或SCAMP 1-YFP融合物的转基因烟草BY-2细胞。共聚焦免疫荧光和免疫金电子显微镜研究表明，YFP-SCAMP 1融合和天然SCAMP 1定位于质膜和移动的结构在转基因BY-2细胞的细胞质中。药物处理和共聚焦免疫荧光研究表明，由YFP-SCAMP 1或SCAMP 1标记的点状细胞器与高尔基体和PVC不同。SCAMP 1标记的细胞器可能代表早期内体，因为内化的内吞标记物FM 4 -64和AM 4 -64在PVC之前到达这些细胞器。此外，渥曼青霉素引起SCAMP 1从早期内体到PVC的重新分布，可能是两个隔室之间融合的结果。高压冷冻/冷冻替代样品的免疫金电子显微镜鉴定SCAMP 1阳性细胞器的管状泡状结构在trans-Golgi与网格蛋白外套。这些早期的内体区室类似于先前描述的植物细胞中的部分包被的网和trans-Golgi网络。
We recently identified multivesicular bodies (MVBs) as prevacuolar compartments (PVCs) in the secretory and endocytic pathways to the lytic vacuole in tobacco ( Nicotiana tabacum) BY-2 cells. Secretory carrier membrane proteins (SCAMPs) are post-Golgi, integral membrane proteins mediating endocytosis in animal cells. To define the endocytic pathway in plants, we cloned the rice ( Oryza sativa) homolog of animal SCAMP1 and generated transgenic tobacco BY-2 cells expressing yellow fluorescent protein (YFP)- SCAMP1 or SCAMP1-YFP fusions. Confocal immunofluorescence and immunogold electron microscopy studies demonstrated that YFP-SCAMP1 fusions and native SCAMP1 localize to the plasma membrane and mobile structures in the cytoplasm of transgenic BY-2 cells. Drug treatments and confocal immunofluorescence studies demonstrated that the punctate cytosolic organelles labeled by YFP-SCAMP1 or SCAMP1 were distinct from the Golgi apparatus and PVCs. SCAMP1-labeled organelles may represent an early endosome because the internalized endocytic markers FM4-64 and AM4-64 reached these organelles before PVCs. In addition, wortmannin caused the redistribution of SCAMP1 from the early endosomes to PVCs, probably as a result of fusions between the two compartments. Immunogold electron microscopy with high-pressure frozen/freeze-substituted samples identified the SCAMP1-positive organelles as tubular-vesicular structures at the trans-Golgi with clathrin coats. These early endosomal compartments resemble the previously described partially coated reticulum and trans-Golgi network in plant cells.