Characterization of the human biliverdin reductase gene structure and regulatory elements: promoter activity is enhanced by hypoxia and suppressed by TNF-α-activated NF-κB
Characterization of the human biliverdin reductase gene structure and regulatory elements: promoter activity is enhanced by hypoxia and suppressed by TNF-α-activated NF-κB
复制标题
DOI：
10.1096/fj.09-144592
复制
发表时间：
2010-09-01
期刊：
影响因子：
4.8
通讯作者：
Maines, Mahin D.
中科院分区：
文献类型：
--
作者：
Gibbs, Peter E. M.;Miralem, Tihomir;Maines, Mahin D.
hBVR是Ser/Thr/Tyr激酶/支架蛋白/转录因子/调节子的细胞内转运蛋白。hBVR是胰岛素/IGF-1/MAPK/PI 3 K信号通路和NF-κ B的上游激活剂。作为还原酶，它将胆绿素转化为抗氧化剂胆红素。hBVR基因有8个外显子，外显子1不翻译。我们报告的hBVR启动子的特性和它的负和正调控，分别由TNF-α和缺氧。通过引物延伸分析确定外显子1的5'端;该外显子上游350-425 bp的抑制剂序列的缺失增强了启动子活性。836 bp启动子中的两个NF-κ B结合位点之一是功能性的; NF-κ B的P65亚基和TNF-α作为抑制剂。基于EMSA和ChIP分析，TNF-α处理增加了NF-κ B与其调节元件的结合。I κ B的过表达增加了hBVR mRNA的表达。胆绿素，而不是胆红素，是有效的抑制hBVR启动子活性的TNF-α。HEK 293 A细胞表达的4种缺氧反应元件（HRE）中只有一种与HIF-1 α和ARNT结合。在HRE的3' G处引入脱碱基位点。该元件在凝胶迁移和细胞内荧光素酶测定中结合HIF-1。缺氧后1、2、4 h，hBVR在细胞核内表达，此时其激酶和还原酶活性均升高。由于缺氧对hBVR启动子和磷酸化有积极影响，TNF-α激活的NF-κ B抑制启动子，而胆绿素抑制NF-κ B活性和hBVR启动子，我们提出了以hBVR/胆绿素为中心的缺氧和TNF-α对NF-κ B的调节机制。吉布斯，体育。M.，Miralem，T.，Maines，M. D.人胆绿素还原酶基因结构和调控元件的表征：缺氧可增强启动子活性，TNF-α激活的NF-κ B可抑制启动子活性。FASEB J.24，3239-3254（2010）。www.fasebj.org
hBVR is a Ser/Thr/Tyr kinase/scaffold protein/transcription factor/intracellular transporter of regulators. hBVR is an upstream activator of the insulin/IGF-1/MAPK/PI3K signaling pathway, and of NF-kappa B. As a reductase, it converts biliverdin to the antioxidant, bilirubin. hBVR gene has 8 exons; exon 1 is not translated. We report the characterization of hBVR promoter and its negative and positive regulation, respectively, by TNF-alpha and hypoxia. The 5' end of exon 1 was defined by primer extension analyses; deletion of an inhibitor sequence 350-425 bp upstream of this exon enhanced the promoter activity. One of two NF-kappa B binding sites in the 836-bp promoter was functional; the P65 subunit of NF-kappa B and TNF-alpha acted as inhibitors. On the basis of EMSA and ChIP assays, TNF-alpha treatment increases binding of NF-kappa B to its regulatory element. Overexpression of I kappa B increased hBVR mRNA. Biliverdin, but not bilirubin, was as effective as TNF-alpha in inhibiting hBVR promoter activity. Only one of 4 hypoxia responsive elements (HREs) bound to HIF-1 alpha and ARNT expressed in HEK293A cells. An abasic site was introduced at the 3' G of the HRE. This element bound HIF-1 in the gel shift and in in-cell luciferase assays. hBVR was detected in the nucleus at 1, 2, and 4 h after hypoxia (1% O-2), at which times its kinase and reductase activities were increased. Because hypoxia positively influences hBVR promoter and phosphorylation and TNF-alpha activated NF-kappa B inhibits the promoter, while biliverdin inhibits both NF-kappa B activity and hBVR promoter, we propose a regulatory mechanism for NF-kappa B by hypoxia and TNF-alpha centered on hBVR/biliverdin.-Gibbs, P. E. M., Miralem, T., Maines, M. D. Characterization of the human biliverdin reductase gene structure and regulatory elements: promoter activity is enhanced by hypoxia and suppressed by TNF-alpha-activated NF-kappa B. FASEB J. 24, 3239-3254 (2010). www.fasebj.org