Jupp R, Hoffmann S, Depto A, Stenberg R M, Ghazal P, Nelson J A
Department of Microbiology and Immunology, Oregon Health Sciences University, Portland 97201.
J Virol. 1993 Sep;67(9):5595-604. doi: 10.1128/JVI.67.9.5595-5604.1993.
The human cytomegalovirus major immediate-early gene encodes several protein isoforms which autoregulate the major immediate-early promoter (MIEP). One of these isoforms, the IE86 protein, represses the MIEP through a DNA sequence located between the TATA box and the transcription initiation site, designated the cis repression signal (crs). Through mutational analysis, amino acid domains within IE86 responsible for binding the crs element were located at the C terminus. Mutation of the putative zinc finger domain, which precluded IE86 from binding DNA, converted the protein from a repressor of MIEP transcription into an activator. DNase I protection analysis demonstrated that the IE86 footprint overlapped the sequence protected by the TATA-binding protein (TBP). Investigation of whether IE86 was able to displace TBP from DNA revealed that both proteins could bind DNA simultaneously. However, higher concentrations of IE86 were required to obtain protection of the crs element in the presence of prebound TBP. Similarly, higher concentrations of TBP were required to obtain protection in the presence of prebound IE86. These observations indicate that steric hinderance impairs but does not prevent both proteins from binding DNA synchronously.
人类巨细胞病毒主要立即早期基因编码几种蛋白质异构体，这些异构体可对主要立即早期启动子（MIEP）进行自我调节。其中一种异构体，即IE86蛋白，通过位于TATA框和转录起始位点之间的一段DNA序列（称为顺式抑制信号，crs）来抑制MIEP。通过突变分析，IE86中负责结合crs元件的氨基酸结构域位于C末端。假定的锌指结构域发生突变，使IE86无法结合DNA，从而将该蛋白从MIEP转录的抑制因子转变为激活因子。DNase I保护分析表明，IE86的足迹与TATA结合蛋白（TBP）保护的序列重叠。对IE86是否能够从DNA上取代TBP的研究表明，两种蛋白可以同时结合DNA。然而，在预先结合TBP的情况下，需要更高浓度的IE86才能获得crs元件的保护。同样，在预先结合IE86的情况下，需要更高浓度的TBP才能获得保护。这些观察结果表明，空间位阻会损害但不会阻止两种蛋白同时结合DNA。
