Schaub M, Krol A, Carbon P
UPR 9002 du CNRS Structure des Macromolécules Biologiques et Mécanismes de Reconnaissance, IBMC, 15 rue René Descartes, 67084 Strasbourg Cedex, France.
J Biol Chem. 1999 Aug 20;274(34):24241-9. doi: 10.1074/jbc.274.34.24241.
The transactivator Staf, which contains seven zinc finger motifs, exerts its effect on gene expression by binding to specific targets in small nuclear RNA (snRNA) and snRNA-type gene promoters. In this work, binding site selection allowed us to identify the 21-base pair ATTACCCATAATGCATYGCGG sequence as the high affinity consensus binding site for Staf. It shows a high sequence divergence with Staf-responsive elements in the Xenopus selenocysteine tRNA (tRNA(Sec)) and human U6 snRNA promoters. By using a combination of approaches, we analyzed the interaction of wild-type and truncated Staf zinc finger domains with the consensus, Xenopus tRNA(Sec), and human U6 sites. Two main conclusions emerged from our data. First, the data clearly indicate that zinc finger 7 does not establish base-specific contacts in Staf-DNA complexes. The second conclusion concerns zinc finger 1, which is required for the binding to the Xenopus tRNA(Sec) site but is dispensable in the case of the human U6 site. Taking into account the sequence differences in the two sites, these findings demonstrate that Staf utilizes zinc finger 1 in a rather flexible manner, illustrating how a protein can interact with DNAs containing targets of different sequences.
反式激活因子Staf含有7个锌指基序，它通过与小核RNA（snRNA）和snRNA型基因启动子中的特定靶点结合来发挥对基因表达的作用。在这项研究中，结合位点筛选使我们能够鉴定出21个碱基对的ATTACCCATAATGCATYGCGG序列作为Staf的高亲和力共有结合位点。它与非洲爪蟾硒代半胱氨酸tRNA（tRNA(Sec)）和人类U6 snRNA启动子中的Staf反应元件存在高度的序列差异。通过使用多种方法相结合，我们分析了野生型和截短的Staf锌指结构域与共有序列、非洲爪蟾tRNA(Sec)和人类U6位点之间的相互作用。我们的数据得出了两个主要结论。首先，数据清楚地表明锌指7在Staf-DNA复合物中不形成碱基特异性接触。第二个结论涉及锌指1，它是与非洲爪蟾tRNA(Sec)位点结合所必需的，但在人类U6位点的情况下是可有可无的。考虑到这两个位点的序列差异，这些发现表明Staf以一种相当灵活的方式利用锌指1，说明了一种蛋白质如何与含有不同序列靶点的DNA相互作用。
