Identification of two internal signal peptide sequences: critical for classical swine fever virus non-structural protein 2 to trans-localize to the endoplasmic reticulum.
Identification of two internal signal peptide sequences: critical for classical swine fever virus non-structural protein 2 to trans-localize to the endoplasmic reticulum.
复制标题
两个内部信号肽序列的鉴定：对于猪瘟病毒非结构蛋白 2 反式定位至内质网至关重要
DOI：
10.1186/1743-422x-8-236
复制
发表时间：
2011-05-18
期刊：
影响因子：
4.8
通讯作者：
He L
中科院分区：
文献类型：
--
作者：
Guo KK;Tang QH;Zhang YM;Kang K;He L
背景猪瘟病毒（classical swine fever virus，CSFV）非结构蛋白2（non-structural 2，NS 2）的膜结构和内质网定位的分子机制尚不清楚。我们试图阐明该蛋白的亚细胞定位，并在我们的研究中负责定位的分子机制。利用逆转录聚合酶链反应扩增NS 2基因，并通过生物信息学分析预测NS 2蛋白的跨膜区和亲水性。采用PCR缺失法扩增了NS 2基因的12个cDNA，并将其克隆到真核表达载体中，转染猪脐静脉内皮细胞系（SUVEC）。NS 2蛋白的亚细胞定位，其特征在于通过共聚焦显微镜，和蛋白质印迹进行分析蛋白质expression.ResultsOur结果表明，-NH 2末端的CSFV NS 2蛋白是高度疏水性和蛋白质定位在ER。至少四个跨膜区和两个内部信号肽序列（氨基酸103 -138和220-262）被鉴定，并认为是其在ER的反式定位的关键。结论本研究首次鉴定了CSFV NS 2蛋白的内部信号肽序列及其亚细胞定位，为进一步研究该蛋白的功能及其在猪瘟病毒致病中的作用奠定了基础。
BackgroundThe membrane topology and molecular mechanisms for endoplasmic reticulum (ER) localization of classical swine fever virus (CSFV) non-structural 2 (NS2) protien is unclear. We attempted to elucidate the subcellular localization, and the molecular mechanisms responsible for the localization of this protein in our study. The NS2 gene was amplified by reverse transcription polymerase chain reaction, with the transmembrane region and hydrophilicity of the NS2 protein was predicted by bioinformatics analysis. Twelve cDNAs of the NS2 gene were amplified by the PCR deletion method and cloned into a eukaryotic expression vector, which was transfected into a swine umbilical vein endothelial cell line (SUVEC). Subcellular localization of the NS2 protein was characterized by confocal microscopy, and western blots were carried out to analyze protein expression.ResultsOur results showed that the -NH2terminal of the CSFV NS2 protein was highly hydrophobic and the protein localized in the ER. At least four transmembrane regions and two internal signal peptide sequences (amino acids103-138 and 220-262) were identified and thought to be critical for its trans-localization to the ER.ConclusionsThis is the first study to identify the internal signal peptide sequences of the CSFV NS2 protein and its subcellular localization, providing the foundation for further exploration of this protein's function of this protein and its role in CSFV pathogenesis.