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首页> 美国卫生研究院文献>Journal of Virology >Interaction of Epstein-Barr Virus Nuclear Antigen Leader Protein (EBNA-LP) with HS1-Associated Protein X-1: Implication of Cytoplasmic Function of EBNA-LP
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Interaction of Epstein-Barr Virus Nuclear Antigen Leader Protein (EBNA-LP) with HS1-Associated Protein X-1: Implication of Cytoplasmic Function of EBNA-LP

机译：爱泼斯坦-巴尔病毒核抗原前导蛋白（EBNA-LP）与HS1相关蛋白X-1的相互作用：EBNA-LP的细胞质功能的暗示。

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Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) consists of W1W2 repeats and a unique C-terminal Y1Y2 domain and has been suggested to play an important role in EBV-induced transformation. To identify the cellular factors interacting with EBNA-LP, we performed a yeast two-hybrid screen, using EBNA-LP cDNA containing four W1W2 repeats as bait and an EBV-transformed human peripheral blood lymphocyte cDNA library as the source of cellular genes. Our results were as follows. (i) All three cDNAs in positive yeast colonies were found to encode the same cellular protein, HS1-associated protein X-1 (HAX-1), which is localized mainly in the cytoplasm and has been suggested to be involved in the regulation of B-cell signal transduction and apoptosis. (ii) Mutational analysis of EBNA-LP revealed that the association with HAX-1 is mediated by the W1W2 repeat domain. (iii) A purified chimeric protein consisting of glutathione S-transferase fused to EBNA-LP specifically formed complexes with HAX-1 transiently expressed in COS-7 cells. (iv) When EBNA-LP and HAX-1 were coexpressed in COS-7 cells, EBNA-LP was specifically coimmunoprecipitated with HAX-1. (v) Careful cell fractionation experiments of an EBV-infected lymphoblastoid cell line revealed that EBNA-LP is localized in the cytoplasm as well as in the nucleus. (vi) When EBNA-LP containing four W1W2 repeats was expressed in COS-7 cells, EBNA-LP was detected mainly in the nucleus by immunofluorescence assay. Interestingly, when EBNA-LP containing a single W1W2 repeat was expressed in COS-7 cells, EBNA-LP was localized predominantly in the cytoplasm and was colocalized with HAX-1. These results indicate that EBNA-LP is in fact present and may have a significant function in the cytoplasm, possibly by interacting with and affecting the function of HAX-1.
机译：爱泼斯坦巴尔病毒（EBV）核抗原前导蛋白（EBNA-LP）由W1W2重复序列和一个独特的C末端Y1Y2结构域组成，已被证明在EBV诱导的转化中起重要作用。为了确定与EBNA-LP相互作用的细胞因子，我们进行了酵母双杂交筛选，使用包含四个W1W2重复序列的EBNA-LP cDNA作为诱饵，并使用EBV转化的人外周血淋巴细胞cDNA文库作为细胞基因的来源。我们的结果如下。 （i）发现在阳性酵母菌落中的所有三个cDNA都编码相同的细胞蛋白HS1相关蛋白X-1（HAX-1），该蛋白主要位于细胞质中，并建议参与其调控。 B细胞信号转导和凋亡。 （ii）EBNA-LP的突变分析表明与HAX-1的缔合是由W1W2重复域介导的。 （iii）由与EBNA-LP融合的谷胱甘肽S-转移酶组成的纯化的嵌合蛋白，与在COS-7细胞中瞬时表达的HAX-1特异性地形成复合物。 （iv）当EBNA-LP和HAX-1在COS-7细胞中共表达时，EBNA-LP与HAX-1特异性共免疫沉淀。 （v）对EBV感染的淋巴母细胞样细胞系进行细心的细胞分级分离实验，发现EBNA-LP位于细胞质以及细胞核中。 （vi）当在COS-7细胞中表达包含四个W1W2重复的EBNA-LP时，通过免疫荧光测定法主要在细胞核中检测到EBNA-LP。有趣的是，当含有单个W1W2重复序列的EBNA-LP在COS-7细胞中表达时，EBNA-LP主要位于细胞质中，并与HAX-1共定位。这些结果表明，实际上存在EBNA-LP，并且可能通过与HAX-1相互作用并影响其功能而在细胞质中具有重要的功能。

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