Identification of a boundary domain adjacent to the potent human cytomegalovirus enhancer that represses transcription of the divergent UL127 promoter
Identification of a boundary domain adjacent to the potent human cytomegalovirus enhancer that represses transcription of the divergent UL127 promoter
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DOI：
10.1128/jvi.74.6.2826-2839.2000
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发表时间：
2000-03-01
影响因子：
5.4
通讯作者：
Ghazal, P
中科院分区：
文献类型：
--
作者：
Angulo, A;Kerry, D;Ghazal, P
复杂的模块化启动子内的转录抑制可能在确定增强子元件的作用中发挥关键作用。在人类巨细胞病毒中？主要立即早期启动子 (MIEP) 基因座包含一个高效且复杂的模块增强子。有证据表明，核苷酸位置 -556 和 -673 之间的 MIEP 序列起到防止来自 UL127 开放阅读框分歧启动子的增强子元件进行转录激活的作用。报告质粒的瞬时转染测定揭示了位于核苷酸-556和-638之间的阻遏序列。使用从含有感染性人巨细胞病毒基因组的细菌人工染色体产生的重组病毒，显示了这些序列在感染情况下赋予抑制的能力。除了-556和-638之间的阻遏序列之外，使用重组病毒突变体的感染实验表明，-638和-673之间的序列也有助于抑制UL127启动子。在体外转录和瞬时转染测定的基础上，我们进一步表明插入的病毒阻遏序列不仅完全抑制增强子介导的同源启动子而且还抑制异源启动子的激活。这些和其他实验表明，抑制涉及宿主编码的调节因子与确定的启动子序列的相互作用，这些启动子序列具有以不依赖染色质的方式近端干扰上游增强子元件的特性。总而言之，我们的研究结果证实了边界域的存在，该边界域可以有效地阻止增强子-启动子相互作用，从而解释增强子如何选择性地激活 MIEP。
Transcriptional repression within a complex modular promoter may play a key role in determining the action of enhancer elements. In human cytomegalovirus? the major immediate-early promoter (MIEP) locus contains a highly potent and complex modular enhancer. Evidence is presented suggesting that sequences of the MIEP between nucleotide positions -556 and -673 function to prevent transcription activation by enhancer elements from the UL127 open reading frame divergent promoter. Transient transfection assays of reporter plasmids revealed repressor sequences located between nucleotides -556 and -638. The ability of these sequences to confer repression in the context of an infection was shown using recombinant viruses generated from a bacterial artificial chromosome containing an infectious human cytomegalovirus genome, In addition to repressor sequences between -556 and -638, infection experiments using recombinant virus mutants indicated that sequences between -638 and -673 also contribute to repression of the UL127 promoter. On the basis of in vitro transcription and transient transfection assays, we further show that interposed viral repressor sequences completely inhibit enhancer-mediated activation of not only the homologous but also heterologous promoters. These and other experiments suggest that repression involves an interaction of host-encoded regulatory factors with defined promoter sequences that have the property of proximally interfering with upstream enhancer elements in a chromatin-independent manner. Altogether, our findings establish the presence of a boundary domain that efficiently blocks enhancer-promoter interactions, thus explaining how the enhancer can work to selectively activate the MIEP.