The adenovirus L4 33-kilodalton protein binds to intragenic sequences of the major late promoter required for late phase-specific stimulation of transcription
The adenovirus L4 33-kilodalton protein binds to intragenic sequences of the major late promoter required for late phase-specific stimulation of transcription
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DOI：
10.1128/jvi.01584-06
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发表时间：
2007-02-01
影响因子：
5.4
通讯作者：
Flint, S. J.
中科院分区：
文献类型：
--
作者：
Ali, Humayra;Leroy, Gary;Flint, S. J.
腺病毒Late Iva(2)蛋白是从病毒主要Late(ML)启动子最有效地转录所必需的，因此，合成大多数病毒Late蛋白是必需的。这种蛋白质是一种序列特异的DNA结合蛋白，也能促进子代病毒颗粒的组装。以前的研究已经证实，IVA(2)蛋白二聚体(DEF-B)与基因内ML启动子序列特异结合，这是后期特异性刺激ML转录所必需的。然而，ML启动子转录的激活与至少一个额外的被称为DEF-A的感染细胞特异性蛋白与启动子的结合有关。利用DEF-A的DNA结合活性测定，我们通过常规纯化方法、标志标记的IVA(2)-蛋白复合物的纯化和病毒晚期蛋白的瞬时合成来鉴定这一未知蛋白。这些实验的结果表明，病毒L433-kDa蛋白是DEF-A的唯一成分：Iva(2)和L433-kDa蛋白是在ML启动子的基因内控制区形成上述所有复合体的必要条件和充分条件。此外，L433-kDa蛋白与启动子结合具有DEF-A的特异性，并在瞬时表达实验中刺激ML启动子的转录。
The adenovirus late IVa(2) protein is required for maximally efficient transcription from the viral major late (ML) promoter, and hence, the synthesis of the majority of viral late proteins. This protein is a sequence-specific DNA-binding protein that also promotes the assembly of progeny virus particles. Previous studies have established that a IVa(2) protein dimer (DEF-B) binds specifically to an intragenic ML promoter sequence necessary for late phase-specific stimulation of ML transcription. However, activation of transcription from the ML promoter correlates with binding of at least one additional infected-cell-specific protein, termed DEF-A, to the promoter. Using an assay for the DNA-binding activity of DEF-A, we identified the unknown protein by using conventional purification methods, purification of FLAG-tagged IVa(2)-protein-containing complexes, and transient synthesis of viral late proteins. The results of these experiments established that the viral L4 33-kDa protein is the only component of DEF-A: the IVa(2) and L4 33-kDa proteins are necessary and sufficient for formation of all previously described complexes in the intragenic control region of the ML promoter. Furthermore, the L4 33-kDa protein binds to the promoter with the specificity characteristic of DEF-A and stimulates transcription from the ML promoter in transient-expression assays.