COOPERATION OF EBV DNA-POLYMERASE AND EA-D(BMRF1) INVITRO AND COLOCALIZATION IN NUCLEI OF INFECTED-CELLS
COOPERATION OF EBV DNA-POLYMERASE AND EA-D(BMRF1) INVITRO AND COLOCALIZATION IN NUCLEI OF INFECTED-CELLS
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DOI：
10.1016/0042-6822(91)90849-7
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发表时间：
1991-09-01
期刊：
影响因子：
3.7
通讯作者：
DORSKY, DI
中科院分区：
文献类型：
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作者：
KIEHL, A;DORSKY, DI
体外转录-翻译表达EB病毒(EBV)DNA聚合酶(EBVpol1)开放阅读框(BALF5)，获得116 kDa的初级翻译产物。酶促DNA聚合酶活性需要47 kDa BMRF1(EA-D)基因产物的存在。在大肠杆菌中表达的BALF5基因产物的抗血清可特异性地在潜伏感染的淋巴母细胞提取液中沉淀出116 kDa的多肽，以诱导EBV复制。免疫荧光显微镜显示EBVpol和EA-D(BMRF1)共定位于诱导细胞的核内离散的病灶；然而，阻断病毒DNA合成导致两种抗原的弥漫性核染色模式。溴脱氧尿嘧啶核苷染色表明，这些与EBVpol共定位的离散病灶是早期病毒DNA合成的部位。这些观察结果表明，EA-D(BMRF1)可能是EBV DNA聚合酶的一种辅助蛋白，它在体内与EBVpol共定位到病毒DNA复制的位置，并在体外协同形成具有活性的EBVpol全酶。
Expression of the Epstein-Barr virus (EBV) DNA polymerase (EBVpol) open reading frame (BALF5) byin vitrotranscription-translation yielded a 116-kDa primary translation product. Enzymatic DNA polymerase activity of thein vitrotranslated polypeptide required the presence of the 47-kDa BMRF1 (EA-D) gene product. Antiserum raised to the BALF5 gene product expressed inEscherichia colispecifically precipitated a 116-kDa polypeptide in extracts of latently infected lymphoblastoid cells induced for EBV replication. Immunofluorescence microscopy revealed colocalization of the EBVpol and EA-D(BMRF1) to discrete foci within the nuclei of induced cells; however, the blockade of viral DNA synthesis resulted in diffuse nuclear staining patterns for both antigens. Bromodeoxyuridine staining of these discrete foci colocalizing with EBVpol suggests that they are sites of early viral DNA synthesis. These observations suggest that EA-D(BMRFl) may be an accessory protein of the EBV DNA polymerase which colocalizesin vivowith EBVpol to sites of viral DNA replication and cooperatesin vitroto form an active EBVpol holoenzyme.