Transcriptional regulation mechanisms of hypoxia-induced neuroglobin gene expression
Transcriptional regulation mechanisms of hypoxia-induced neuroglobin gene expression
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DOI：
10.1042/bj20111856
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发表时间：
2012-04-01
影响因子：
4.1
通讯作者：
Wang, Xiaoying
中科院分区：
文献类型：
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作者：
Liu, Ning;Yu, Zhanyang;Wang, Xiaoying
Ngb（神经红蛋白）已被确定为一种新型内源性神经保护剂。然而，人们对 Ngb 表达的调控机制知之甚少，尤其是在缺氧条件下。在本研究中，我们将小鼠 Ngb 基因的核心近端启动子定位到 554 bp 片段，该片段包含假定保守的 NF-kappa B（核因子 kappa B）和 Egr（早期生长反应因子 1）结合位点。转录因子 p65、p50、Egr1 或 Sp1（特异性蛋白 1）的过表达和敲低分别会增加和减少 Ngb 表达。对突变 Ngb 基因启动子构建体转染的实验评估以及 EMSA（电泳迁移率变化测定）和 ChIP（染色质免疫沉淀）测定表明，NF-kappa B 家族成员（p65、p50 等！）、Egr1 和 Sp1 在体外和体内与 Ngb 基因的近端启动子区域结合。此外，还发现 kappa B3 位点是负责缺氧诱导的 Ngb 启动子活性的关键顺式元件。 NF-kappa B (p65) 和 Sp1 也与缺氧诱导的 Ngb 表达上调有关。尽管小鼠Ngb基因的启动子中没有保守的HRE（缺氧反应元件），但本研究的结果表明HIF-1α（缺氧诱导因子1α）也参与缺氧诱导的Ngb上调。总之，我们发现 NF-kappa B、Egr1 和 Sp1 通过与小鼠 Ngb 启动子的特异性相互作用，在基础 Ngb 表达的调节中发挥重要作用。 NF-kappa B、Sp1 和 HIF-1 α 有助于缺氧条件下小鼠 Ngb 基因表达的上调。
Ngb (neuroglobin) has been identified as a novel endogenous neuroprotectant. However, little is known about the regulatory mechanisms of Ngb expression, especially under conditions of hypoxia. In the present study, we located the core proximal promoter of the mouse Ngb gene to a 554 bp segment, which harbours putative conserved NF-kappa B (nuclear factor kappa B)- and Egr (early growth-response factor 1) -binding sites. Overexpression and knockdown of transcription factors p65, p50, Egr1 or Sp1 (specificity protein 1) increased and decreased Ngb expression respectively. Experimental assessments with transfections of mutational Ngb gene promoter constructs, as well as EMSA (electrophoretic mobility-shift assay) and ChIP (chromatin immunoprecipitation) assays, demonstrated that NF-kappa B family members (p65, p50 and cite!), Egr1 and Sp1 bound in vitro and in vivo to the proximal promoter region of the Ngb gene. Moreover, a kappa B3 site was found as a pivotal cis-element responsible for hypoxia-induced Ngb promoter activity. NF-kappa B (p65) and Sp1 were also responsible for hypoxia-induced up-regulation of Ngb expression. Although there are no conserved HREs (hypoxia-response elements) in the promoter of the mouse Ngb gene, the results of the present study suggest that HIF-1 alpha (hypoxia-inducible factor-1 alpha) is also involved in hypoxia-induced Ngb up-regulation. In conclusion, we have identified that NF-kappa B, Egr1 and Sp1 played important roles in the regulation of basal Ngb expression via specific interactions with the mouse Ngb promoter. NF-kappa B, Sp1 and HIF-1 alpha contributed to the up-regulation of mouse Ngb gene expression under hypoxic conditions.