The THAP domain of THAP1 is a large C2CH module with zinc-dependent sequence-specific DNA-binding activity
The THAP domain of THAP1 is a large C2CH module with zinc-dependent sequence-specific DNA-binding activity
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DOI：
10.1073/pnas.0406882102
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发表时间：
2005-05-10
影响因子：
11.1
通讯作者：
Girard, JP
中科院分区：
文献类型：
--
作者：
Clouaire, T;Roussigne, M;Girard, JP
我们最近描述了一种进化上保守的蛋白质基序，命名为THAP结构域，它定义了一个以前没有特征的细胞因子家族（TRAP蛋白）。THAP结构域表现出与果蝇P元件转座酶的位点特异性DNA结合结构域的相似性，包括推定的金属配位C2CH签名（CX 2 - 4CX 35 - 53 CX 2 H）。在这篇文章中，我们报告了一个全面的清单，约100个不同的THAP蛋白在模型动物生物体，包括人类核促凋亡因子THAP 1和DAP 4/THAP 0，转录抑制因子THAP 7，斑马鱼的直系同源细胞周期调节因子E2 F6，和秀丽隐杆线虫染色质相关蛋白HIM-17和细胞周期调节因子LIN-36和LIN-15 B。此外，我们证明了THAP结构域作为锌指超家族的锌依赖性序列特异性DNA结合结构域的生化功能。在体外结合位点的选择，使我们能够确定一个11个核苷酸的共识DNA结合序列，特异性识别的TRAP结构域的人THAP 1。该序列中的单核苷酸位置的突变废除了TRAP结构域结合。用锌螯合剂1，10-邻菲咯啉的实验表明THAP结构域是锌依赖性DNA结合结构域。单半胱氨酸或组氨酸残基的定点突变支持的锌协调和DNA结合活性的C2CH基序的作用。另外四个保守的残基（P，W，F和P），定义THAP的共有序列，也被发现是DNA结合所需的。结合前人在C. elegans，我们的研究结果表明，细胞TRAP蛋白可能作为锌依赖性序列特异性DNA结合因子的增殖，凋亡，细胞周期，染色体分离，染色质修饰和转录调控的作用。
We have recently described an evolutionarily conserved protein motif, designated the THAP domain, which defines a previously uncharacterized family of cellular factors (TRAP proteins). The THAP domain exhibits similarities to the site-specific DNA-binding domain of Drosophila P element transposase, including a putative metal-coordinating C2CH signature (CX2-4CX35-53CX2H). In this article, we report a comprehensive list of approximate to 100 distinct THAP proteins in model animal organisms, including human nuclear proapoptotic factors THAP1 and DAP4/THAP0, transcriptional repressor THAP7, zebrafish orthologue of cell cycle regulator E2F6, and Caenorhabditis elegans chromatin-associated protein HIM-17 and cell-cycle regulators LIN-36 and LIN-15B. In addition, we demonstrate the biochemical function of the THAP domain as a zinc-dependent sequence-specific DNA-binding domain belonging to the zinc-finger superfamily. In vitro binding-site selection allowed us to identify an 11-nucleotide consensus DNA-binding sequence specifically recognized by the TRAP domain of human THAP1. Mutations of single nucleotide positions in this sequence abrogated TRAP-domain binding. Experiments with the zinc chelator 1,10-o-phenanthroline revealed that the THAP domain is a zinc-dependent DNA-binding domain. Site-directed mutagenesis of single cysteine or histidine residues supported a role for the C2CH motif in zinc coordination and DNA-binding activity. The four other conserved residues (P, W, F, and P), which define the THAP consensus sequence, were also found to be required for DNA binding. Together with previous genetic data obtained in C. elegans, our results suggest that cellular TRAP proteins may function as zinc-dependent sequence-specific DNA-binding factors with roles in proliferation, apoptosis, cell cycle, chromosome segregation, chromatin modification, and transcriptional regulation.