Farina Antonella, Cardinali Giorgia, Santarelli Roberta, Gonnella Roberta, Webster-Cyriaque Jennifer, Bei Roberto, Muraro Raffaella, Frati Luigi, Angeloni Antonio, Torrisi Maria Rosaria, Faggioni Alberto
Istituto Pasteur Fondazione Cenci Bolognetti, Dipartimento di Medicina Sperimentale e Patologia, Università La Sapienza, Rome, Italy.
J Med Virol. 2004 Jan;72(1):102-11. doi: 10.1002/jmv.10561.
A novel protein encoded by the BFRF1 gene of the Epstein-Barr virus was identified recently [Farina et al. (2000) J Virol 74:3235-3244], which is antigenic "in vivo" and expressed early in the viral replicative cycle. In the present study, its subcellular localization was examined in greater detail comparing Epstein-Barr virus (EBV) induced producing and nonproducing cell lines by immunofluorescence: in 12-0-tetradecanoyl phorbol-13-acetate (TPA)-induced Raji and B95-8 cells, as well as in anti-IgG-stimulated Akata cells, the protein appeared to be localized over the cell nuclear membrane. A similar nuclear membrane localization was observed in epithelial cells of oral hairy leukoplakia, a pathological manifestation of permissive EBV infection. In contrast, upon transfection of BFRF1 in the EBV-negative Burkitt's lymphoma cell line DG75, the protein was localized predominantly over the plasma membrane. The membrane localization was abolished when DG75 cells were transfected with a C-terminal deletion mutant of BFRF1 lacking the transmembrane domain. Because induced Raji cells do not produce virus, the above observations indicate that the nuclear membrane localization is not associated with viral production, but requires the expression of EBV genes, and suggest that additional proteins, expressed early during viral lytic infection, might be necessary to target the protein to the nuclear membrane. Immunogold electron microscopy on ultrathin cryosections of induced B95-8 cells showed that BFRF1 on the nuclear membranes was concentrated over multilayered domains representing areas of active viral replication or at the sites of viral budding, suggesting that BFRF1 is involved in the process of viral assembly.
最近发现了一种由爱泼斯坦-巴尔病毒（EBV）的BFRF1基因编码的新型蛋白质[法里纳等人（2000年），《病毒学杂志》74:3235 - 3244]，它在“体内”具有抗原性，并且在病毒复制周期早期表达。在本研究中，通过免疫荧光比较EBV诱导的产生病毒和不产生病毒的细胞系，对其亚细胞定位进行了更详细的研究：在12 - O - 十四烷酰佛波醇 - 13 - 乙酸酯（TPA）诱导的拉吉细胞和B95 - 8细胞以及抗IgG刺激的阿卡塔细胞中，该蛋白质似乎定位于细胞核膜上。在允许EBV感染的病理表现——口腔毛状白斑的上皮细胞中也观察到了类似的核膜定位。相比之下，将BFRF1转染到EBV阴性的伯基特淋巴瘤细胞系DG75中时，该蛋白质主要定位于质膜上。当用缺乏跨膜结构域的BFRF1的C末端缺失突变体转染DG75细胞时，膜定位被消除。由于诱导的拉吉细胞不产生病毒，上述观察结果表明核膜定位与病毒产生无关，但需要EBV基因的表达，并提示在病毒裂解感染早期表达的其他蛋白质可能是将该蛋白质靶向核膜所必需的。对诱导的B95 - 8细胞超薄冷冻切片进行免疫金电子显微镜观察显示，核膜上的BFRF1集中在代表活跃病毒复制区域的多层结构域上或病毒出芽部位，这表明BFRF1参与病毒组装过程。
