Identification of a novel protein encoded by the BamHI A region of the Epstein-Barr virus
Identification of a novel protein encoded by the BamHI A region of the Epstein-Barr virus
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DOI：
10.1128/jvi.71.4.2765-2771.1997
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发表时间：
1997-04-01
影响因子：
5.4
通讯作者：
RaabTraub, N
中科院分区：
文献类型：
--
作者：
Fries, KL;Sculley, TB;RaabTraub, N
先前已在鼻咽癌 (NPC) 组织中发现了一组通过 BamHI A 片段转录的不寻常的 3' 共端剪接 mRNA。在伯基特淋巴瘤 (BL) 细胞系和 Epstein-Barr 病毒 (EBV) 转化的淋巴细胞中也检测到了低水平的这些 RNA。对来自 C15 NPC 肿瘤的 cDNA 文库的克隆进行序列分析表明，初级转录本进行了差异剪接，产生了一系列相关转录本，所有这些转录本都包含转录本 3' 端的 BARF0 开放阅读框 (ORF)。鉴定出一种 cDNA 在 5' 端延伸 BARF0 ORF，形成 RK-BARF0 ORF。在这项研究中，制备了针对代表由 BARF0 ORF 编码的氨基酸序列的合成肽的兔抗血清。该抗血清检测到谷胱甘肽 S-转移酶-BARF0 融合蛋白以及从 H1299 细胞中的转染构建体表达的 BARF0 和 RK-BARF0 蛋白。血清还免疫沉淀 20-kDa BARF0 和 30-kDa RK-BARF0 体外翻译蛋白。免疫印迹分析在所有测试的 EBV 感染细胞系中发现了 30 和 35 kDa 的蛋白质双联体。细胞分级分离研究表明这些蛋白质是膜相关的。在细胞系中检测到的蛋白质的大小及其与膜的关联表明它们可能由 RK-BARF0 转录本编码，预计该转录本包含膜定位信号。在 NPC 活检和 BL 活检制备的蛋白质提取物中也检测到了这些蛋白质，但在毛状白斑（一种允许的 EBV 感染）中未检测到。这些结果表明，来自 EBV BamHI A 区域的向右 RNA 转录本编码一种或多种在潜伏感染细胞和肿瘤组织中表达的蛋白质。
An unusual set of 3' coterminal, spliced mRNAs transcribed through the BamHI A fragment have been previously identified in nasopharyngeal carcinoma (NPC) tissues. These RNAs have also been detected at low levels in Burkitt's lymphoma (BL) cell lines and Epstein-Barr virus (EBV)-transformed lymphocytes. Sequence analyses of clones from a cDNA library derived from the C15 NPC tumor indicated that the primary transcripts are differentially spliced, giving rise to a family of related transcripts, all of which encompass the BARF0 open reading frame (ORF) at the 3' end of the transcripts. One cDNA was identified that extended the BARF0 ORF at the 5' end, forming the RK-BARF0 ORF. In this study, a rabbit antiserum to a synthetic peptide representing an amino acid sequence encoded by the BARF0 ORF was prepared. This antiserum detected a glutathione S-transferase-BARF0 fusion protein and both BARF0 and RK-BARF0 proteins expressed from transfected constructs in H1299 cells. The serum also immunoprecipitated the 20-kDa BARF0 and 30-kDa RK-BARF0 in vitro-translated proteins. Immunoblot analyses identified a protein doublet of 30 and 35 kDa in all of the EBV-infected cell lines tested. Cellular fractionation studies revealed that the proteins were membrane associated. The sizes of the proteins detected in cell lines and their association with membranes suggests that they are likely encoded by the RK-BARF0 transcript, which is predicted to contain a membrane localization signal. The proteins were also detected in protein extracts prepared from NPC biopsies and a BL biopsy but not from hairy leukoplakia, a permissive EBV infection. These results reveal that the rightward RNA transcripts from the BamHI A region of EBV encode one or more proteins that are expressed in latently infected cells and in tumor tissue.