Isolation and functional characterization of TIF-IB, a factor that confers promoter specificity to mouse RNA polymerase I.
Isolation and functional characterization of TIF-IB, a factor that confers promoter specificity to mouse RNA polymerase I.
复制标题
TIF-IB 的分离和功能表征，TIF-IB 是赋予小鼠 RNA 聚合酶 I 启动子特异性的因子。
DOI：
10.1093/nar/18.6.1385
复制
发表时间：
1990
影响因子：
14.9
通讯作者：
I. Grummt
中科院分区：
文献类型：
--
作者：
A. Schnapp;J. Clos;W. Hädelt;R. Schreck;A. Cvekl;I. Grummt
鼠核糖体基因启动子含有两个顺式作用控制元件，它们协同作用以促进 RNA 聚合酶 I 有效且准确的转录起始。起始位点近端核心元件是 RNA 聚合酶 I (pol I) 启动子识别所必需的，涵盖位置 -39 至 -1 的序列。位于核苷酸-142和-112之间的上游控制元件(UCE)可刺激体内和体外转录起始的效率。在此，我们报告了特定 rDNA 结合蛋白（转录起始因子 TIF-IB）的分离和功能表征，该蛋白与小鼠核糖体 RNA 基因启动子的核心区域特异性相互作用。高度纯化的 TIF-IB 在其他两种必需起始因子 TIF-IA 和 TIF-IC 存在的情况下补充转录活性。我们证明，UCE 的顺式存在大大增强了纯化的 TIF-IB 与核心启动子的结合效率。在整个纯化过程中观察到上游序列对 TIF-IB 结合的这种积极作用，表明两个远距离启动子元件的协同作用不是由不同于 TIF-IB 的蛋白质介导的。增加两个控制元件之间的距离仍然有利于稳定的因子结合，但消除了转录激活。结果表明，TIF-IB 与 rDNA 启动子的结合是功能性转录起始复合物组装中必不可少的早期步骤。然而，TIF-IB 随后与其他辅助转录起始因子的相互作用需要 UCE 和核心启动子元件之间的正确间距。
The murine ribosomal gene promoter contains two cis-acting control elements which operate in concert to promote efficient and accurate transcription initiation by RNA polymerase I. The start site proximal core element which is indispensable for promoter recognition by RNA polymerase I (pol I) encompasses sequences from position -39 to -1. An upstream control element (UCE) which is located between nucleotides -142 and -112 stimulates the efficiency of transcription initiation both in vivo and in vitro. Here we report the isolation and functional characterization of a specific rDNA binding protein, the transcription initiation factor TIF-IB, which specifically interacts with the core region of the mouse ribosomal RNA gene promoter. Highly purified TIF-IB complements transcriptional activity in the presence of two other essential initiation factors TIF-IA and TIF-IC. We demonstrate that the binding efficiency of purified TIF-IB to the core promoter is strongly enhanced by the presence in cis of the UCE. This positive effect of upstream sequences on TIF-IB binding is observed throughout the purification procedure suggesting that the synergistic action of the two distant promoter elements is not mediated by a protein different from TIF-IB. Increasing the distance between both control elements still facilitates stable factor binding but eliminates transcriptional activation. The results demonstrate that TIF-IB binding to the rDNA promoter is an essential early step in the assembly of a functional transcription initiation complex. The subsequent interaction of TIF-IB with other auxiliary transcription initiation factors, however, requires the correct spacing between the UCE and the core promoter element.