Hypoxia-induced synthesis of hemoglobin in the crustacean Daphnia magna is hypoxia-inducible factor-dependent
Hypoxia-induced synthesis of hemoglobin in the crustacean Daphnia magna is hypoxia-inducible factor-dependent
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DOI：
10.1074/jbc.m403981200
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发表时间：
2004-08-20
影响因子：
4.8
通讯作者：
Bunn, HF
中科院分区：
文献类型：
--
作者：
Gorr, TA;Cahn, JD;Bunn, HF
在淡水甲壳类动物大水蚤（Daphnia magna）中存在的四种已知的珠蛋白基因中，有几种是由缺氧单独诱导的，在缺氧的情况下，苍白的常温动物会呈现出明显的红色。水蚤珠蛋白基因的启动子区域包含许多缺氧反应元件（HREs），作为缺氧诱导因子（hif）的潜在结合位点。与人类HRE序列结合的水蚤HIF在缺氧（红色）动物的提取物中检测到，而在常氧（苍白）动物的提取物中检测不到。利用系统发育保守的HIF/HRE识别，我们采用异种转染表达HIF的人和果蝇细胞来模拟水蚤的HIF信号传导。这些实验表明，在D. magna球蛋白-2基因启动子内的三个功能性HREs协同作用，最大限度地缺氧诱导下游荧光素酶报告基因。这三个顺式元件中的两个，在启动子位置-258和-107，能够在体外特异性结合人、果蝇或水蚤的HIF复合物。在低氧张力条件下，在内源性人或过表达果蝇HIF蛋白存在的情况下，这两个位点也是最大限度地诱导报告转录所必需的。球蛋白-2基因启动子的第三个基序，位于-146位的CACGTG回文，作为未知组成转录因子的对接位点。在人类细胞中，这种-146复合物干扰HIF在邻近-107 HRE的占用，从而控制HIF介导的下游靶标缺氧激活的程度。
Of the four known globin genes that exist in the freshwater crustacean Daphnia magna, several are individually induced by hypoxia, lending pale normoxic animals a visible red color when challenged by oxygen deprivation. The promoter regions of the Daphnia globin genes each contain numerous hypoxia response elements (HREs) as potential binding sites for hypoxia-inducible factors (HIFs). Daphnia HIF, bound to human HRE sequences, was detected in extracts from hypoxic (red), but not normoxic (pale), animals. Taking advantage of the phylogenetically conserved HIF/HRE recognition, we employed heterologous transfections of HIF-expressing human and Drosophila cells to model HIF signaling in Daphnia. These experiments revealed that three functional HREs within the promoter of the D. magna globin-2 gene cooperate for maximal hypoxic induction of a downstream luciferase reporter gene. Two of these three cis-elements, at promoter positions -258 and -107, are able to specifically bind human, Drosophila, or Daphnia HIF complexes in vitro. The same two sites are also necessary for maximal induction of reporter transcription under low oxygen tension in the presence of either endogenous human or overexpressed Drosophila HIF proteins. The third motif of the globin-2 gene promoter, a CACGTG palindrome at position -146, functions as a docking site for an unknown constitutive transcription factor. In human cells, this -146 complex interferes with HIF occupancy at the adjacent -107 HRE and thus controls the extent of HIF-mediated hypoxic activation of the downstream target.