Schweiger Michal-Ruth, You Jianxin, Howley Peter M
Department of Pathology, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, Massachusetts 02115, USA.
J Virol. 2006 May;80(9):4276-85. doi: 10.1128/JVI.80.9.4276-4285.2006.
The papillomavirus E2 regulatory protein has essential roles in viral transcription and the initiation of viral DNA replication as well as for viral genome maintenance. Brd4 has recently been identified as a major E2-interacting protein and, in the case of the bovine papillomavirus type 1, serves to tether E2 and the viral genomes to mitotic chromosomes in dividing cells, thus ensuring viral genome maintenance. We have explored the possibility that Brd4 is involved in other E2 functions. By analyzing the binding of Brd4 to a series of alanine-scanning substitution mutants of the human papillomavirus type 16 E2 N-terminal transactivation domain, we found that amino acids required for Brd4 binding were also required for transcriptional activation but not for viral DNA replication. Functional studies of cells expressing either the C-terminal domain of Brd4 that can bind E2 and compete its binding to Brd4 or short interfering RNA to knock down Brd4 protein levels revealed a role for Brd4 in the transcriptional activation function of E2 but not for its viral DNA replication function. Therefore, these studies establish a broader role for Brd4 in the papillomavirus life cycle than as the chromosome tether for E2 during mitosis.
乳头瘤病毒E2调节蛋白在病毒转录、病毒DNA复制起始以及病毒基因组维持过程中发挥着重要作用。Brd4最近被鉴定为一种主要的与E2相互作用的蛋白，在1型牛乳头瘤病毒中，它能将E2和病毒基因组与分裂细胞中的有丝分裂染色体相连，从而确保病毒基因组的维持。我们探究了Brd4参与其他E2功能的可能性。通过分析Brd4与人乳头瘤病毒16型E2 N端反式激活结构域的一系列丙氨酸扫描取代突变体的结合情况，我们发现Brd4结合所需的氨基酸对于转录激活是必需的，但对于病毒DNA复制并非必需。对表达能够结合E2并竞争其与Brd4结合的Brd4 C端结构域的细胞，或对敲低Brd4蛋白水平的短发夹RNA进行功能研究，结果显示Brd4在E2的转录激活功能中发挥作用，但在其病毒DNA复制功能中并非如此。因此，这些研究表明Brd4在乳头瘤病毒生命周期中的作用比其在有丝分裂期间作为E2的染色体连接蛋白更为广泛。
