Localizing the movement proteins of Abutilon mosaic geminivirus in yeast by subcellular fractionation and freeze-fracture immuno-labelling
Localizing the movement proteins of Abutilon mosaic geminivirus in yeast by subcellular fractionation and freeze-fracture immuno-labelling
复制标题
DOI：
10.1007/s705-002-8305-8
复制
发表时间：
2002-01-01
影响因子：
2.7
通讯作者：
Jeske, H
中科院分区：
文献类型：
--
作者：
Aberle, HJ;Rütz, ML;Jeske, H
利用裂殖酵母（Schizosophylomycespombe）细胞表达了与谷胱甘肽-S-转移酶（GST）和Flag-peptide融合的Abutilonmosaicgeminivirus运动蛋白BC 1和BV 1，分析了这些蛋白在缺乏植物特异性因子的真核细胞中的基本胞内分布。经细胞裂解和差速离心后，BC 1蛋白质大部分迅速沉淀。使用冷冻断裂免疫标记，蛋白质原位检测主要在质膜和较低程度上在细胞质囊泡，但不在细胞质，细胞核，或线粒体。抗BC 1，抗GST和抗Flag抗体标记的光滑florescent仅在质膜的原生质面。BC 1的行为，其使用在酵母双杂交分析的后果进行了讨论。相反，BV 1主要在细胞核中检测到，部分在细胞质中，但从未与膜。
The movement proteins BC1 and BV1 of Abutilon mosaic geminivirus fused to glutathion-S-transferase (GST) and Flag-peptide were expressed in fission yeast (Schizosaccharomyces pombe) cells to analyse the fundamental intracellular distribution of these proteins in an eukaryotic cell in the absence of plant-specific factors. Most of BC1 protein sedimented rapidly after cell lysis and differential centrifugation. Using freeze-fracture immuno-labelling, the protein was detected in situ predominantly at plasma membranes and to a lower extent at cytoplasmic vesicles but not in the cytoplasm, the nuclei, or the mitochondria. Anti-BC1, anti-GST, and anti-Flag antibodies tagged smooth flecks only at the protoplasmic faces of the plasma membrane. The consequences of the BC1 behaviour for its use in two-hybrid analysis in yeast are discussed. In contrast, BV1 was detected mainly in the nucleus and partially in the cytoplasm but never associated with membranes.