Differential transcriptional regulation of the two vascular endothelial growth factor receptor genes - Flt-1, but not Flk-1/KDR, is up-regulated by hypoxia
Differential transcriptional regulation of the two vascular endothelial growth factor receptor genes - Flt-1, but not Flk-1/KDR, is up-regulated by hypoxia
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DOI：
10.1074/jbc.272.38.23659
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发表时间：
1997-09-19
影响因子：
4.8
通讯作者：
Ferrara, N
中科院分区：
文献类型：
--
作者：
Gerber, HP;Condorelli, F;Ferrara, N
血管内皮生长因子（VEGF）及其两种内皮细胞特异性受体酪氨酸激酶Flk-1/KDR和Flt-1在生理和病理性血管生成中发挥关键作用。缺氧已被证明是体内 VEGF 及其受体上调的主要机制。当我们将人脐静脉内皮细胞暴露于体外缺氧条件时，我们观察到Flt-1表达水平增加，相反，Flk-1/KDR mRNA水平没有变化或轻微抑制，这些发现表明缺氧对两种受体的转录调节存在差异，为了鉴定参与缺氧反应的调节元件，分离了小鼠Flt-1和Flk-1/KDR基因的启动子区域 并与荧光素酶报告基因结合进行测试。在瞬时转染测定中，缺氧导致 Flt-1 启动子强烈转录激活，而 Flk-1/KDR 转录基本没有变化，启动子缺失分析表明 Flt-1 启动子的 430 bp 区域是缺氧反应转录激活所必需的，该区域包含与先前发现的缺氧诱导因子 1 (HIF) 共有结合位点相匹配的七聚体序列 在其他缺氧诱导基因，如 VEGF 基因和促红细胞生成素基因中，我们进一步将介导缺氧反应的元件范围缩小到 40 碱基对序列，包括假定的 HIF 结合位点。 缺氧导致的转录激活受损。这些发现表明，与 KDR/Flk-1 基因不同，Flt-1 受体基因通过位于 Flt-1 启动子位置 -976 至 -937 的缺氧诱导增强子元件直接因缺氧而上调。
Vascular endothelial growth factor (VEGF) and its two endothelial cell-specific receptor tyrosine kinases, Flk-1/KDR and Flt-1, play a key role in physiological and pathological angiogenesis. Hypoxia has been shown tee be a major mechanism for up-regulation of VEGF and its receptors in vivo. When we exposed human umbilical vein endothelial cells to hypoxic conditions in vitro, we observed increased levels of Flt-1 expression, In contrast, Flk-1/KDR mRNA levels were unchanged or slightly repressed, These findings suggest a differential transcriptional regulation of the two receptors by hypoxia, To identify regulatory elements involved in the hypoxic response, promoter regions of the mouse Flt-1 and Flk-1/KDR genes were isolated and tested in conjunction with luciferase reporter gene. In transient transfection assays, hypoxia led to strong transcriptional activation of the Flt-1 promoter, whereas Flk-1/KDR transcription was essentially unchanged, Promoter deletion analysis demonstrated a 430-bp region of the Flt-1 promoter to be required for transcriptional activation in response to hypoxia, This region includes a heptamer sequence matching the hypoxia-inducible factor-1 (HIF) consensus binding site previously found in other hypoxia-inducible genes such as the VEGF gene and erythropoietin gene, We further narrowed down the element mediating the hypoxia response to a 40-base pair sequence including the putative HIF binding site, We show that this element acts like an enhancer, since it activated transcription irrespective of its location or orientation in the construct, Furthermore, mutations within the putative HIF consensus binding site lead to impaired transcriptional activation by hypoxia, These findings indicate that, unlike the KDR/Flk-1 gene, the Flt-1 receptor gene is directly up-regulated by hypoxia via a hypoxia-inducible enhancer element located at positions -976 to -937 of the Flt-1 promoter.