The KRAB zinc finger gene ZNF74 encodes an RNA-binding protein tightly associated with the nuclear matrix
The KRAB zinc finger gene ZNF74 encodes an RNA-binding protein tightly associated with the nuclear matrix
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DOI：
10.1074/jbc.271.26.15458
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发表时间：
1996-06-28
影响因子：
4.8
通讯作者：
Aubry, M
中科院分区：
文献类型：
--
作者：
Grondin, B;Bazinet, M;Aubry, M
我们之前克隆了 ZNF74，这是一种在 DiGeorge 综合征中通常缺失的发育表达锌指基因。这里展示了人类基因的内含子/外显子组织和表达蛋白质的功能特性。来自转录因子 IIIA/Kruppel 家族的锌指基因包含三个外显子。位于预测的 64 kDa 锌指蛋白 N 末端的截短 Kruppel 相关盒 (KRAB) 由外显子 2 编码。包括锌指结构域和 3'-非翻译区 (UTR) 在内的蛋白质其余部分由外显子 3 编码。5'-UTR（外显子 1）和 3'-UTR 均包含重复的 Alu 元件。编码整个 ZNF74 编码区的 cDNA 的体外翻译产生了 63-kDa 的蛋白质，如在十二烷基硫酸钠-聚丙烯酰胺凝胶上测定的。使用显示与 (65) 锌紧密结合的细菌表达的融合蛋白来测试 ZNF74 的核酸结合特性。通过 RNA 结合测定，发现 ZNF74 与聚 (U) 和聚 (G) RNA 均聚物特异性结合。与这些均聚物的有限结合而不是与聚 (A) 和聚 (C) 的结合表明 ZNF74 显示出 RNA 序列偏好。 RNA 结合由锌指结构域介导。对转染细胞的免疫荧光研究揭示了 ZNF74 核定位。在细胞核中观察到的标记模式清楚地排除了核仁。锌指区域缺乏经典的核定位信号，但被发现负责核靶向，亚细胞和原位连续分级进一步表明 ZNF74 与核基质相关。该蛋白的 RNA 结合特性及其与核基质（参与 DNA 复制以及 RNA 合成和加工的亚核区室）的紧密联系表明 ZNF74 在 RNA 代谢中的作用。
We previously cloned ZNF74, a developmentally expressed zinc finger gene commonly deleted in DiGeorge syndrome. Here, the intron/exon organization of the human gene and the functional properties of the expressed protein are presented. This zinc finger gene from the transcription factor IIIA/Kruppel family contains three exons. A truncated Kruppel-associated box (KRAB) located at the N terminus of the predicted 64-kDa zinc finger protein is encoded by exon 2. The remainder of the protein including the zinc finger domain as well as the 3'-untranslated region (UTR) is encoded by exon 3. Both 5'-UTR (exon 1) and 3'-UTR contain repetitive Alu elements. In vitro translation of a cDNA encoding the entire ZNF74 coding region produced a 63-kDa protein as determined on sodium dodecyl sulfate-polyacrylamide gel. A bacterially expressed fusion protein shown to bind tightly to (65)zinc was used to test the nucleic acid binding properties of ZNF74. By RNA binding assays, ZNF74 was found to bind specifically to poly(U) and poly(G) RNA homopolymers. The restricted binding to these homopolymers and not to poly(A) and poly(C) suggested that ZNF74 displays RNA sequence preferences. RNA binding was mediated by the zinc finger domain. Immunofluorescence studies on transfected cells revealed ZNF74 nuclear localization. The labeling pattern observed in the nuclei clearly excluded the nucleoli. The zinc finger region lacks a classical nuclear localization signal but was found to be responsible for nuclear targeting, Subcellular and in situ sequential fractionations further showed that ZNF74 is associated with the nuclear matrix. The RNA binding properties of this protein and its tight association with the nuclear matrix, a subnuclear compartment involved in DNA replication as well as RNA synthesis and processing, suggest a role for ZNF74 in RNA metabolism.