Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia.
Expression of Iron Regulatory Protein 1 Is Regulated not only by HIF-1 but also pCREB under Hypoxia.
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缺氧条件下铁调节蛋白1的表达不仅受到HIF-1的调节，还受到pCREB的调节
DOI：
10.7150/ijbs.16437
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发表时间：
2016
影响因子：
9.2
通讯作者：
Ke Y
中科院分区：
文献类型：
--
作者：
Luo QQ;Qian ZM;Zhou YF;Zhang MW;Wang D;Zhu L;Ke Y
IRP1和HIF-1α对缺氧反应的不一致以及IRP1和pCREB含量变化的相似趋势使我们推测pCREB可能参与了缺氧条件下IRP1的调节。在这里，我们使用定量 PCR、蛋白质印迹、免疫荧光、电泳迁移率变动分析 (EMSA) 和染色质免疫沉淀 (ChIP) 研究了缺氧下 pCREB 在 HepG2 细胞中 IRP1 表达中的作用。我们证明了 1) 缺氧会增加细胞核内的 pCREB 水平； 2) 在IRP1基因中发现了推定的CRE； 3）缺氧处理的HepG2细胞核提取物可以与CRE1和CRE3结合，而假定的CRE的100倍竞争剂可以不同程度地消除结合活性； 4) EMSA的CRE1和CRE3 DNA-蛋白质复合物中发现pCREB； 5) IRP1的CRE1和CRE3结合活性取决于CREB激活而不是HIF-1； 6) LY294002可以阻止缺氧条件下IRP1表达增加； 7) ChIP 实验证明 pCREB 与 IRP1 启动子结合； 8)HIF-1和/或HIF-2 siRNA对缺氧8小时的细胞中pCREB和IRP1蛋白的表达没有影响。我们的研究结果证明了 pCREB 参与了 IRP1 表达，并揭示了 PI3K/Akt 通路在缺氧条件下 CREB 激活中的主导作用，还表明缺氧条件下 HIF-1 和 pCERB 或其他转录因子对 IRP1 表达的双重调节可能是大多数（如果不是全部）缺氧诱导基因的共同机制。
The inconsistent of responses of IRP1 and HIF-1 alpha to hypoxia and the similar tendencies in the changes of IRP1 and pCREB contents led us to hypothesize that pCREB might be involved in the regulation of IRP1 under hypoxia. Here, we investigated the role of pCREB in IRP1 expression in HepG2 cells under hypoxia using quantitative PCR, western blot, immunofluorescence, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). We demonstrated that 1) Hypoxia increased pCREB levels inside of the nucleus; 2) Putative CREs were found in the IRP1 gene; 3) Nuclear extracts of HepG2 cells treated with hypoxia could bind to CRE1 and CRE3, and 100-fold competitor of putative CREs could abolish the binding activity to varying degrees; 4) pCREB was found in the CRE1 and CRE3 DNA-protein complexes of EMSA; 5) CRE1 and CRE3 binding activity of IRP1 depended on CREB activation but not on HIF-1; 6) Increased IRP1 expression under hypoxia could be prevented by LY294002; 7) ChIP assays demonstrated that pCREB binds to IRP1 promoter; and 8) HIF-1 and/or HIF-2 siRNA had no effect on the expression of pCREB and IRP1 proteins in cells treated with hypoxia for 8 hours. Our findings evidenced for the involvement of pCREB in IRP1 expression and revealed a dominant role of PI3K/Akt pathway in CREB activation under hypoxia and also suggested that dual-regulation of IRP1 expression by HIF-1 and pCERB or other transcription factor(s) under hypoxia might be a common mechanism in most if not all of hypoxia-inducible genes.
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DOI：
10.4049/jimmunol.1101532
发表时间：
2011-12-01
期刊：
Journal of immunology (Baltimore, Md. : 1950)
影响因子：
--
作者：
Li X;Han X;Llano J;Bole M;Zhou X;Swan K;Anandaiah A;Nelson B;Patel NR;Reinach PS;Koziel H;Tachado SD
通讯作者：
Tachado SD
影响因子：
82.9
作者：
Donnelly, SC;Haslett, C;Bucala, R
通讯作者：
Bucala, R
影响因子：
4.8
作者：
Hanson, ES;Foot, LM;Leibold, EA
通讯作者：
Leibold, EA
影响因子：
4.7
作者：
Hara, T;Hamada, J;Ushio, Y
通讯作者：
Ushio, Y
影响因子：
4.7
作者：
Lee, Hsueh-Te;Chang, Ying-Chao;Huang, Chao-Ching
通讯作者：
Huang, Chao-Ching