Seibl R, Motz M, Wolf H
J Virol. 1986 Dec;60(3):902-9. doi: 10.1128/JVI.60.3.902-909.1986.
The expression of the 1,800-base-pair BamHI Z region of Epstein-Barr virus DNA was analyzed by hybrid-selected translation with several DNA subclones and RNA from different cell lines. Furthermore, large segments of the three reading frames extending in this area were expressed as fusion proteins into Escherichia coli. The fusion proteins were partially purified and used to immunize rabbits. These sera were used to confirm our mapping assignments and to identify the respective posttranslationally modified proteins in in vivo labeling experiments. The reading frame BRLF1 (the first reading frame starting in the BamHI R fragment in leftward orientation) encoded a 93- to 96-kilodalton (kDa) protein depending on the cell line. The molecular weight of in vivo-labeled proteins was increased relative to that of in vitro-translated proteins, indicating that a posttranslational modification had occurred. The BZLF1 reading frame encoded a 35-kDa protein. It was posttranslationally cleaved from a 38-kDa precursor in induced B95-8 and induced Raji cells and from a 40-kDa precursor in induced P3HR1 cells. In Raji cells superinfected with virus derived from P3HR1 cells, the protein seemed to be expressed both from endogenous Raji genomes and from infecting genomes. The transcripts for the 93- to 96-kDa and the 35-kDa protein overlapped partially. The serum against the expressed third reading frame BZLF2 specifically precipitated a 140-kDa protein. This reading frame contains only 650 nucleotides, and therefore further coding sequences were presumably spliced to BZLF2. The latter is deleted in the Raji cell line; therefore, the observed 140kDa protein in superinfected Raji cells was expressed from infecting P3HR1 genomes.
通过与几个DNA亚克隆以及来自不同细胞系的RNA进行杂交选择翻译，分析了爱泼斯坦-巴尔病毒DNA的1800个碱基对的BamHI Z区域的表达。此外，在该区域延伸的三个阅读框的大片段被表达为融合蛋白进入大肠杆菌。融合蛋白被部分纯化并用于免疫兔子。这些血清用于确认我们的定位任务，并在体内标记实验中鉴定各自翻译后修饰的蛋白质。阅读框BRLF1（以向左方向从BamHI R片段开始的第一个阅读框）根据细胞系的不同，编码一种93至96千道尔顿（kDa）的蛋白质。相对于体外翻译的蛋白质，体内标记蛋白质的分子量增加，表明发生了翻译后修饰。BZLF1阅读框编码一种35 kDa的蛋白质。它在诱导的B95-8细胞和诱导的Raji细胞中从38 kDa的前体经翻译后切割产生，在诱导的P3HR1细胞中从40 kDa的前体经翻译后切割产生。在被源自P3HR1细胞的病毒超感染的Raji细胞中，该蛋白质似乎既从内源性Raji基因组表达，也从感染的基因组表达。93至96 kDa和35 kDa蛋白质的转录本部分重叠。针对表达的第三个阅读框BZLF2的血清特异性沉淀出一种140 kDa的蛋白质。这个阅读框仅包含650个核苷酸，因此推测进一步的编码序列被剪接到BZLF2上。后者在Raji细胞系中缺失；因此，在超感染的Raji细胞中观察到的140 kDa蛋白质是从感染的P3HR1基因组表达的。
