Distinct roles of two binding sites for the bovine papillomavirus (BPV) E2 transactivator on BPV DNA replication.
Distinct roles of two binding sites for the bovine papillomavirus (BPV) E2 transactivator on BPV DNA replication.
复制标题
牛乳头瘤病毒 (BPV) E2 反式激活子的两个结合位点对 BPV DNA 复制的不同作用。
DOI：
10.1128/jvi.72.7.5735-5744.1998
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发表时间：
1998
影响因子：
5.4
通讯作者：
Borowiec,JA
中科院分区：
文献类型：
--
作者：
Gillette,TG;Borowiec,JA
用牛乳头瘤病毒1型（BPV）研究了转录因子对DNA复制的调节作用。BPV在体内的复制需要两种病毒蛋白：起始结合蛋白E1和转录反激活子E2。在起源中，E1与一个中心区域相互作用，中心区域两侧是E2的两个结合位点（BS11和BS12），其中只有BS12被报道为体内复制所必需的。通过化学干扰和电泳迁移转移实验，我们发现E2与每个位点的结合刺激了不同e1起源复合物的形成。利用临界E2触点与BS12和E1触点与双对称元形成高迁移率的C1配合物。相反，E2与另一侧起始侧的BS11元件的相互作用促进了低迁移率C3复合物的形成。C3是一种类似于C2的新物种，C2是一种由E1单独形成的具有复制活性的复合体。E1在C1和C3复合物中的结合有很大的不同，C1复合物中的E1局限于原点双对称区，而C3复合物中的E1则包括从BS11近缘到BS12远缘的区域。我们发现e2结合位点的存在对于体内野生型复制活性以及C3复合体的最大生产是必要的。这些结果表明，在正常的病毒环境中，BS11和BS12在病毒DNA复制的启动中发挥独立但协同的作用，这取决于它们在起源中的位置。我们的数据表明，E2与每个位点的结合依次刺激不同的e1起源复合物的形成，导致复制能力复合物的形成。
The modulation of DNA replication by transcription factors was examined by using bovine papillomavirus type 1 (BPV). BPV replication in vivo requires two viral proteins: E1, an origin-binding protein, and E2, a transcriptional transactivator. In the origin, E1 interacts with a central region flanked by two binding sites for E2 (BS11 and BS12), of which only BS12 has been reported to be essential for replication in vivo. Using chemical interference and electrophoretic mobility shift assays, we found that the binding of E2 to each site stimulates the formation of distinct E1-origin complexes. A high-mobility C1 complex is formed by using critical E2 contacts to BS12 and E1 contacts to the dyad symmetry element. In contrast, interaction of E2 with the BS11 element on the other origin flank promotes the formation of the lower-mobility C3 complex. C3 is a novel species that resembles C2, a previously identified complex that is replication active and formed by E1 alone. The binding of E1 greatly differs in the C1 and C3 complexes, with E1 in the C1 complex limited to the origin dyad symmetry region and E1 in the C3 complex encompassing the region from the proximal edge of BS11 through the distal edge of BS12. We found that the presence of both E2-binding sites is necessary for wild-type replication activity in vivo, as well as for maximal production of the C3 complex. These results show that in the normal viral context, BS11 and BS12 play separate but synergetic roles in the initiation of viral DNA replication that are dependent on their location within the origin. Our data suggest a model in which the binding of E2 to each site sequentially stimulates the formation of distinct E1-origin complexes, leading to the replication-competent complex.