Interaction of the Vaccinia Virus RNA Polymerase-Associated 94-Kilodalton Protein with the Early Transcription Factor
Interaction of the Vaccinia Virus RNA Polymerase-Associated 94-Kilodalton Protein with the Early Transcription Factor
复制标题
DOI：
10.1128/jvi.01653-09
复制
发表时间：
2009-12-01
影响因子：
5.4
通讯作者：
Moss, Bernard
中科院分区：
文献类型：
--
作者：
Yang, Zhilong;Moss, Bernard
痘苗病毒（VACV）编码的多亚基RNA聚合酶（RPO）与特定因子结合，转录早期、中期和晚期病毒基因。然而，另一种被称为 94 kDa RPO 相关蛋白 (RAP94) 的病毒编码多肽与 RPO 紧密结合，用于早期基因的转录。与八个 RPO 核心亚基不同，RAP94 仅在感染后后期合成。此外，RAP94 对于包装 RPO 和组装病毒颗粒的早期转录所需的其他组件是必需的。 RAP94 与 NPH I（转录终止所需的 DNA 依赖性 ATP 酶）和多功能 Poly(A) 聚合酶小亚基/2'-O-甲基转移酶/延伸因子的直接关联已被证明。人们曾怀疑 RAP94 在核心 RPO 和 VACV 早期转录因子 (VETF) 之间提供结构和功能联系，但之前并未得到证实。使用组成型或诱导性表达 VETF 亚基和带有亲和标签的 RAP94 的 VACV 重组体，我们表明（i）VETF 在体内和体外仅与含有 RAP94 的 RPO 结合，（ii）RAP94 与 VETF 的结合需要后者的两个亚基，（iii）RAP94 与 VETF 和核心 RPO 亚基的相互作用不需要病毒 DNA 或其他病毒编码的晚期蛋白，（iv） RAP94的不同结构域结合VETF和RPO的核心亚基，并且(v)NPH I和VETF独立且可能同时结合至RAP94的N端区域。因此，RAP94 在 RPO 和转录起始、延伸和终止所需的蛋白质之间提供了桥梁。
A multisubunit RNA polymerase (RPO) encoded by vaccinia virus (VACV), in conjunction with specific factors, transcribes early, intermediate, and late viral genes. However, an additional virus-encoded polypeptide referred to as the RPO-associated protein of 94 kDa (RAP94) is tightly bound to the RPO for the transcription of early genes. Unlike the eight RPO core subunits, RAP94 is synthesized exclusively at late times after infection. Furthermore, RAP94 is necessary for the packaging of RPO and other components needed for early transcription in assembling virus particles. The direct association of RAP94 with NPH I, a DNA-dependent ATPase required for transcription termination, and the multifunctional poly(A) polymerase small subunit/2'-O-methyltransferase/ elongation factor was previously demonstrated. That RAP94 provides a structural and functional link between the core RPO and the VACV early transcription factor (VETF) has been suspected but not previously demonstrated. Using VACV recombinants that constitutively or inducibly express VETF subunits and RAP94 with affinity tags, we showed that (i) VETF associates only with RPO containing RAP94 in vivo and in vitro, (ii) the association of RAP94 with VETF requires both subunits of the latter, (iii) neither viral DNA nor other virus-encoded late proteins are required for the interaction of RAP94 with VETF and core RPO subunits, (iv) different domains of RAP94 bind VETF and core subunits of RPO, and (v) NPH I and VETF bind independently and possibly simultaneously to the N-terminal region of RAP94. Thus, RAP94 provides the bridge between the RPO and proteins needed for transcription initiation, elongation, and termination.