Camenisch G, Stroka D M, Gassmann M, Wenger R H
Institute of Physiology, University of Zürich-Irchel, 8057 Zürich, Switzerland.
Pflugers Arch. 2001 Nov;443(2):240-9. doi: 10.1007/s004240100679.
Hypoxia-inducible factors (HIFs) locate to HIF-binding sites (HBSs) within the hypoxia-response elements (HREs) of oxygen-regulated genes. Whereas HIF-1alpha is expressed ubiquitously, HIF-2alpha is found primarily in the endothelium, similar to endothelin-1 (ET-1) and fms-like tyrosine kinase 1 (Flt-1), the expression of which is controlled by HREs. We identified an unique sequence alteration in both ET-1 and Flt-1 HBSs not found in other HIF-1 target genes, implying that these HBSs might cause binding of HIF-2 rather than HIF-1. However, electrophoretic mobility shift assays showed HIF-1 and HIF-2 DNA complex formation with the unique ET-1 HBS to be about equal. Both DNA-binding and hypoxic activation of reporter genes using the ET-1 HBS was decreased compared with transferrin and erythropoietin HBSs. The Flt-1 HBS was non-functional when assayed in isolation, suggesting that additional factors are required for hypoxic up-regulation via the reported Flt-1 HRE. Interestingly, HIF-1 activity could be restored fully by point-mutating the ET-1 (but not the Flt-1) HBS, suggesting that the wild-type ET-1 HBS attenuated the full hypoxic response known from other oxygen-regulated genes. Such a mechanism might serve to limit the expression of this potent vasoconstrictor in hypoxia.
缺氧诱导因子（HIFs）定位于氧调节基因的缺氧反应元件（HREs）内的HIF结合位点（HBSs）。虽然HIF-1α在全身广泛表达，但HIF-2α主要在内皮细胞中发现，类似于内皮素-1（ET-1）和fms样酪氨酸激酶1（Flt-1），它们的表达受HREs控制。我们在ET-1和Flt-1的HBSs中发现了一种独特的序列改变，而在其他HIF-1靶基因中未发现，这意味着这些HBSs可能导致HIF-2而非HIF-1的结合。然而，电泳迁移率变动分析表明，HIF-1和HIF-2与独特的ET-1 HBS形成的DNA复合物数量大致相等。与转铁蛋白和促红细胞生成素的HBSs相比，使用ET-1 HBS的报告基因的DNA结合和缺氧激活均降低。单独检测时，Flt-1 HBS无功能，这表明通过报道的Flt-1 HRE进行缺氧上调还需要其他因素。有趣的是，通过对ET-1（而非Flt-1）HBS进行点突变可完全恢复HIF-1活性，这表明野生型ET-1 HBS减弱了其他氧调节基因已知的完全缺氧反应。这种机制可能有助于限制这种强效血管收缩剂在缺氧时的表达。
