Transcriptional activation of the human leptin gene in response to hypoxia - Involvement of hypoxia-inducible factor 1
Transcriptional activation of the human leptin gene in response to hypoxia - Involvement of hypoxia-inducible factor 1
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DOI：
10.1074/jbc.m205172200
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发表时间：
2002-09-13
影响因子：
4.8
通讯作者：
Flores-Riveros, J
中科院分区：
文献类型：
--
作者：
Ambrosini, G;Nath, AK;Flores-Riveros, J
除了在能量稳态中发挥重要作用外，瘦素还作为一种具有多种生理效应功能的多效细胞因子而出现。最近发现的瘦素的促血管生成活性表明了这样的假设：瘦素的产生可能受到缺氧的调节，其他血管生成因子也是如此。为了检验这一提议，测量了瘦素蛋白和 mRNA 的表达，发现瘦素蛋白和 mRNA 的表达在环境或化学缺氧（暴露于去铁胺或氯化钴后）中显着上调，这种效应需要完整的 RNA 合成，这表明转录机制。用与报告基因连接的瘦素基因启动子的缺失构建体瞬时转染培养细胞，揭示了位于近端上游区域内的-116位的功能性缺氧反应元件（HRE）。这种假定的 HRE 具有特征性 5'-RCGTG-3' 核心基序，这是缺氧敏感基因的标志，并被缺氧诱导因子 1 (HIF1) 识别，HIF1 由 HIF1α/HIFβ 异二聚体组成。含有这种 -116/HRE 的构建体支持报告基因在缺氧时的表达，但在突变时则不然。常氧细胞中 HIF1α cDNA 的表达模拟了用野生型瘦素 -116/HRE 构建体共转染的细胞中缺氧诱导的报告基因表达，但与突变型瘦素 -116/HRE 构建体共转染的细胞不同。使用 P-32 标记的瘦素启动子 -116/HRE 探针和缺氧处理细胞的核提取物进行的凝胶位移测定表明 HIF1α/β 异二聚体的结合，该结合被过量的未标记 -116/HRE 探针或来自促红细胞生成素基因增强子的 HIF1 结合探针阻断。总而言之，这些观察结果表明，瘦素基因通过缺氧敏感基因常用的转录途径与缺氧积极参与。
In addition to having a major role in energy homeostasis, leptin is emerging as a pleiotropic cytokine with multiple physiological effector functions. The recently discovered proangiogenic activity of leptin suggested the hypothesis that its production might be regulated by hypoxia, as are other angiogenic factors. To examine this proposal, the expression of leptin protein and mRNA was measured and found to be markedly up-regulated in response to ambient or chemical hypoxia (upon exposure to desferrioxamine or cobalt chloride), an effect that requires intact RNA synthesis, suggesting a transcriptional mechanism. Transient transfection of cultured cells with deletion constructs of the leptin gene promoter linked to a reporter gene revealed a functional hypoxia response element (HRE) located at position -116 within the proximal upstream region. This putative HRE harbors a characteristic 5'-RCGTG-3' core motif, a hallmark of hypoxia-sensitive genes and recognized by the hypoxia-inducible factor 1 (HIF1), which consists of a HIF1alpha/HIFbeta heterodimer. Constructs harboring this -116/HRE supported reporter gene expression in response to hypoxia but not when mutated. Expression of HIF1alpha cDNA in normoxic cells mimicked hypoxia-induced reporter gene expression in cells cotransfected with the wild type leptin -116/HRE construct but not with the mutant. Gel shift assays with a P-32-labeled leptin promoter -116/HRE probe and nuclear extracts from hypoxia-treated cells indicated binding of the HIF1alpha/beta heterodimer, which was blocked with an excess of unlabeled -116/HRE probe or a HIF1-binding probe from the erythropoietin gene enhancer. Taken together, these observations demonstrate that the leptin gene is actively engaged by hypoxia through a transcriptional pathway commonly utilized by hypoxia-sensitive genes.