FUNCTIONAL DISSECTION OF THE AUTOGRAPHA-CALIFORNICA NUCLEAR POLYHEDROSIS-VIRUS ENHANCER ELEMENT HR5
FUNCTIONAL DISSECTION OF THE AUTOGRAPHA-CALIFORNICA NUCLEAR POLYHEDROSIS-VIRUS ENHANCER ELEMENT HR5
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DOI：
10.1006/viro.1994.1197
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发表时间：
1994-05-01
期刊：
影响因子：
3.7
通讯作者：
DONG, W
中科院分区：
文献类型：
--
作者：
GUARINO, LA;DONG, W
苜蓿银纹夜蛾核型多角体病毒的hr 5增强子元件在病毒反式激活因子IE 1存在下刺激杆状病毒延迟早期启动子的转录。凝胶阻滞分析的hr 5片段和提取物之间的相互作用，从细胞转染IE 1编码质粒揭示了三个DNA-蛋白质复合物的存在。为了更好地确定hr 5增强子的功能域，我们构建了一组增强子元件中含有部分缺失的质粒。在瞬时测定中测试这些构建体的体外DNA结合活性和增强子功能。结果表明，DNA-蛋白质相互作用所需的最小序列是保守的24-bp回文序列的一半，包含在一个60-bp的直接重复序列（DR 60）。然而，增强子功能所需的最小序列是DR 60的完整拷贝。模板挑战实验表明，IE 1与DR 60的完整或半拷贝以相等的亲和力结合。缺失分析通过使用合成寡核苷酸的体外结合和瞬时表达测定来证实。(C)1994年出版社出版。
The hr5 enhancer element of Autographa californica nuclear polyhedrosis virus stimulates transcription from baculovirus-delayed early promoters in the presence of the viral transactivator, IE1. Gel retardation analyses of interactions between a fragment of hr5 and extracts prepared from cells transfected with an IE1-encoding plasmid revealed the presence of three DNA-protein complexes. In order to better define the functional domains of the hr5 enhancer, we constructed a set of plasmids containing partial deletions in the enhancer element. These constructs were tested for both in vitro DNA binding activity and enhancer function in transient assays. The results indicated that the minimum sequence required for DNA-protein interactions was half of the conserved 24-bp palindrome that is contained within a 60-bp direct repeat (DR60). However, the minimum sequence required for enhancer function was a complete copy of DR60. Template challenge experiments indicated that IE1 bound with equal affinity with a complete or a half copy of DR60. The deletion analyses were confirmed by in vitro binding and transient expression assays with synthetic oligonucleotides. (C) 1994 Academic Press, Inc.