Application of green fluorescent protein-labeled assay for the study of subcellular localization of Newcastle disease virus matrix protein
Application of green fluorescent protein-labeled assay for the study of subcellular localization of Newcastle disease virus matrix protein
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DOI：
10.1016/j.jviromet.2013.08.014
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发表时间：
2013-12-01
影响因子：
3.1
通讯作者：
Liu, Xiufan
中科院分区：
文献类型：
--
作者：
Duan, Zhiqiang;Li, Qunhui;Liu, Xiufan
绿色荧光蛋白（GFP）作为一种强有力的体内基因表达标记物，已广泛应用于研究活细胞中蛋白质的定位和功能。本研究采用绿色荧光蛋白（GFP）标记的方法，对不同毒力和基因型的纽卡斯尔病毒（NDV）毒株的基质（M）蛋白进行了亚细胞定位研究。10株NDV的M蛋白与GFP融合蛋白（GFP-M）在整个转染过程中均定位于细胞核和核仁中，而GFP-M蛋白在整个转染过程中均定位于细胞核和核仁中。另外两株在转染早期在细胞核和核仁中表达，晚期在细胞质中表达。此外，还研究了GFP-M融合蛋白中先前定义的核定位信号的突变。位置262和263处的单个变化不影响M的核定位，而将这两个精氨酸残基改变为天冬酰胺引起M主要重新定位于细胞质。GFP-M是研究M蛋白亚细胞定位的合适系统，可用于进一步鉴定负责M蛋白核仁定位和胞质定位的信号序列。(C)2013爱思唯尔有限公司版权所有。
Green fluorescent protein (GFP) used as a powerful marker of gene expression in vivo has so far been applied widely in studying the localizations and functions of protein in living cells. In this study, GFP-labeled assay was used to investigate the subcellular localization of matrix (M) protein of different virulence and genotype Newcastle disease virus (NDV) strains. The M protein of ten NDV strains fused with GFP (GFP-M) all showed nuclear-and-nucleolar localization throughout transfection, whereas that. of the other two strains were observed in the nucleus and nucleolus early in transfection but in the cytoplasm late in transfection. In addition, mutations to the previously defined nuclear localization signal in the GFP-M fusion protein were studied as well. Single changes at positions 262 and 263 did not affect nuclear localization of M, while changing both of these arginine residues to asparagine caused re-localization of M mainly to the cytoplasm. The GFP-M was validated as a suitable system for studying the subcellular localization of M protein and could be used to assist us in further identifying the signal sequences responsible for the nucleolar localization and cytoplasmic localization of M protein. (C) 2013 Elsevier B.V. All rights reserved.