Identification of a putative transactivation domain in human Nanog
Identification of a putative transactivation domain in human Nanog
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DOI：
10.1038/emm.2005.33
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发表时间：
2005-06-30
影响因子：
12.8
通讯作者：
Kim, JH
中科院分区：
文献类型：
--
作者：
Oh, JH;Do, HJ;Kim, JH
Nanog是新近发现的一种分化的同源结构域蛋白，可指导胚胎干细胞的无限繁殖并维持其多能性。据报道，小鼠Nanog在N端和C端有两个有效的反式激活结构域。人Nanog(HNanog)多肽与小鼠Nanog的开放阅读框和同源结构域的同源性分别为58%和87%。然而，人们对hNanog的功能结构域和分子机制知之甚少。在这项研究中，我们首次提出只有hNanog的C末端包含一个有效的反式激活结构域。根据Homeobox结构域的氨基酸序列，我们将hNanog开放阅读框大致划分为N端、同源域和C端三个区域，构建了hNanog个体与GaI4 DNA结合域的融合蛋白或天然hNanog蛋白的上下文。使用含有GaI4或Nanog结合位点的报告载体进行的报告分析显示，唯一的C-末端区域显示出显著的反式激活折叠诱导。然而，有趣的是，与小鼠Nanog不同，N-末端区域没有显著的激活作用，这表明C-末端区域可能在靶基因的转录激活中发挥更关键的作用。综上所述，hNanog中一个可能的反式激活结构域的发现可能有助于进一步了解参与人类干细胞自我更新和多能性的下游基因调控的分子机制。
Nanog is a newly identified divergent homeodomain protein that directs the infinite propagation and sustains the pluripotency of embryonic stem cells. It has been reported that murine Nanog has two potent transactivation domains in N-terminal and C-terminal regions. Human Nanog (hNanog) polypeptide shares about 58% and 87% identity to the open reading frame and homeodomain of murine Nanog, respectively. However, the functional domains and molecular mechanisms of hNanog are poorly understood. In this study, for the first time, we presented that only C-terminus of hNanog contains a potent transactivation domain. Based on the amino acid sequences of homeobox domain, we roughly divided hNanog open reading frame into the three regions such as N-terminal, homeodomain and C-terminal regions and constructed either the fusion proteins between hNanog individual and GaI4 DNA binding domain or the context of native hNanog protein. Reporter assays by using reporter plamid containing GaI4 or Nanog binding site revealed that the only C-terminal region exhibited the significant fold induction of transactivation. However, interestingly, there was no significant activation through N-terminal region unlike murine Nanog, suggesting that C-terminal region may have more critical roles in the transcriptional activation of target genes. Taken, together, the finding of a putative transactivation, domain in hNanog may contribute to the further understanding of molecular mechanism on the regulation of downstream genes involved in self-renewal and pluripotency of human stem cells.