Subcellular localization of the 30K protein in TMV-inoculated tobacco protoplasts.
Subcellular localization of the 30K protein in TMV-inoculated tobacco protoplasts.
复制标题
接种 TMV 的烟草原生质体中 30K 蛋白的亚细胞定位。
DOI：
10.1016/0042-6822(86)90143-1
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发表时间：
1986
期刊：
影响因子：
3.7
通讯作者：
Y. Okada
中科院分区：
文献类型：
--
作者：
Y. Watanabe;I. Ooshika;T. Meshi;Y. Okada
我们利用[35S]蛋氨酸的脉冲标记和脉冲追踪实验，研究了30K蛋白在tmv接种的烟草原生质体中的胞内定位。原生质体用非离子洗涤剂裂解，提取液离心得到可溶性和粗核馏分。30K蛋白大部分存在于粗核部分。核馏分进一步纯化在逐步珀科尔梯度离心。细胞核和30K蛋白在相同的馏分中发现。脉冲追踪实验结果表明，30K蛋白在可溶性部分合成，然后转移到粗核部分。Ls1的30K蛋白是一种温度敏感（ts）突变体，影响细胞间病毒转运功能，即使在不允许的温度下，也在细胞核中发现。这些结果表明，30K蛋白必须定位在细胞核中才能发挥作用，30K蛋白向细胞核的易位受损并不是突变体Ls1的原因。
We investigated the intracellular localization of the 30K protein in TMV-inoculated tobacco protoplasts by means of pulse-labeling and pulse-chase experiments with [35S]methionine. Protoplasts were lysed with a nonionic detergent and the extracts were centrifuged to yield soluble and crude nuclear fractions. Most of the 30K protein was found in the crude nuclear fraction. The nuclear fraction was further purified by centrifugation in a step-wise Percoll gradient. Nuclei and the 30K protein were found in the same fractions. The results of pulse-chase experiments indicated that the 30K protein is synthesized in the soluble fraction and then translocated to the crude nuclear fraction. The 30K protein of Ls1, a temperature-sensitive (ts) mutant affecting the cell-to-cell viral transport function, was also found in the nuclei, even at a nonpermissive temperature. These results suggested that the 30K protein has to be localized in the nuclei to function, and that impaired translocation of the 30K protein to the nuclei is not responsible for thetslesion of mutant Ls1.