Department of Life Sciences and Institute of Genome Sciences, National Yang-Ming University, Taipei 11221, Taiwan.
J Biol Chem. 2010 Apr 2;285(14):10189-97. doi: 10.1074/jbc.M109.049130. Epub 2010 Feb 4.
The mammalian embryonic zeta-globin genes, including that of humans, are expressed at the early embryonic stage and then switched off during erythroid development. This autonomous silencing of the zeta-globin gene transcription is probably regulated by the cooperative work of various protein-DNA and protein-protein complexes formed at the zeta-globin promoter and its upstream enhancer (HS-40). We present data here indicating that a protein-binding motif, ZF2, contributes to the repression of the HS-40-regulated human zeta-promoter activity in erythroid cell lines and in transgenic mice. Combined site-directed mutagenesis and EMSA suggest that repression of the human zeta-globin promoter is mediated through binding of the zinc finger factor RREB1 to ZF2. This model is further supported by the observation that human zeta-globin gene transcription is elevated in the human erythroid K562 cell line or the primary erythroid culture upon RNA interference (RNAi)(2) knockdown of RREB1 expression. These data together suggest that RREB1 is a putative repressor for the silencing of the mammalian zeta-globin genes during erythroid development. Because zeta-globin is a powerful inhibitor of HbS polymerization, our experiments have provided a foundation for therapeutic up-regulation of zeta-globin gene expression in patients with severe hemoglobinopathies.
哺乳动物胚胎 ζ-珠蛋白基因，包括人类的 ζ-珠蛋白基因，在胚胎早期表达，然后在红细胞发育过程中关闭。 ζ-珠蛋白基因转录的这种自主沉默可能是由 ζ-珠蛋白启动子及其上游增强子 (HS-40) 上形成的各种蛋白质-DNA 和蛋白质-蛋白质复合物的协同作用调节的。我们在这里提供的数据表明，一个蛋白结合基序 ZF2 有助于抑制红细胞谱系细胞系和转基因小鼠中 HS-40 调节的人 ζ-启动子活性。联合定点诱变和 EMSA 表明，通过锌指因子 RREB1 与 ZF2 的结合来介导人 ζ-珠蛋白启动子的抑制。该模型进一步得到以下观察结果的支持：在人红细胞 K562 细胞系或原代红细胞培养物中，通过 RNA 干扰 (RNAi)(2) 敲低 RREB1 表达，人 ζ-珠蛋白基因转录升高。这些数据共同表明，RREB1 是哺乳动物 ζ-珠蛋白基因在红细胞发育过程中沉默的潜在抑制剂。因为 ζ-珠蛋白是 HbS 聚合的有力抑制剂，我们的实验为严重血红蛋白病患者中 ζ-珠蛋白基因表达的治疗性上调提供了基础。
