Wellmann Sven, Bührer Christoph, Moderegger Eva, Zelmer Andrea, Kirschner Renate, Koehne Petra, Fujita Jun, Seeger Karl
Department of Pediatric Oncology/Hematology, Charité Campus Virchow-Klinikum, Medical University of Berlin, 13353 Berlin, Germany.
J Cell Sci. 2004 Apr 1;117(Pt 9):1785-94. doi: 10.1242/jcs.01026. Epub 2004 Mar 16.
The transcriptional regulation of several dozen genes in response to low oxygen tension is mediated by hypoxia-inducible factor 1 (HIF-1), a heterodimeric protein composed of two subunits, HIF-1alpha and HIF-1beta. In the HIF-1alpha-deficient human leukemic cell line, Z-33, exposed to mild (8% O(2)) or severe (1% O(2)) hypoxia, we found significant upregulation of two related heterogenous nuclear ribonucleoproteins, RNA-binding motif protein 3 (RBM3) and cold inducible RNA-binding protein (CIRP), which are highly conserved cold stress proteins with RNA-binding properties. Hypoxia also induced upregulation of RBM3 and CIRP in the murine HIF-1beta-deficient cell line, Hepa-1 c4. In various HIF-1 competent cells, RBM3 and CIRP were induced by moderate hypothermia (32 degrees C) but hypothermia was ineffective in increasing HIF-1alpha or vascular endothelial growth factor (VEGF), a known HIF-1 target. In contrast, iron chelators induced VEGF but not RBM3 or CIRP. The RBM3 and CIRP mRNA increase after hypoxia was inhibited by actinomycin-D, and in vitro nuclear run-on assays demonstrated specific increases in RBM3 and CIRP mRNA after hypoxia, which suggests that regulation takes place at the level of gene transcription. Hypoxia-induced RBM3 or CIRP transcription was inhibited by the respiratory chain inhibitors NaN(3) and cyanide in a dose-dependent fashion. However, cells depleted of mitochondria were still able to upregulate RBM3 and CIRP in response to hypoxia. Thus, RBM3 and CIRP are adaptatively expressed in response to hypoxia by a mechanism that involves neither HIF-1 nor mitochondria.
缺氧诱导因子1（HIF-1）介导了几十种基因对低氧张力的转录调控，HIF-1是一种异二聚体蛋白，由两个亚基HIF-1α和HIF-1β组成。在缺乏HIF-1α的人白血病细胞系Z-33中，分别暴露于轻度（8% O₂）或重度（1% O₂）缺氧环境下，我们发现两种相关的异质性核核糖核蛋白——RNA结合基序蛋白3（RBM3）和冷诱导RNA结合蛋白（CIRP）显著上调，它们是具有RNA结合特性的高度保守的冷应激蛋白。缺氧也能诱导小鼠缺乏HIF-1β的细胞系Hepa-1 c4中RBM3和CIRP的上调。在各种具有HIF-1活性的细胞中，适度低温（32℃）可诱导RBM3和CIRP，但低温对增加HIF-1α或血管内皮生长因子（VEGF，一种已知的HIF-1靶标）无效。相反，铁螯合剂可诱导VEGF，但不能诱导RBM3或CIRP。缺氧后RBM3和CIRP mRNA的增加被放线菌素-D抑制，体外核转录分析表明缺氧后RBM3和CIRP mRNA有特异性增加，这表明调控发生在基因转录水平。缺氧诱导的RBM3或CIRP转录被呼吸链抑制剂NaN₃和氰化物以剂量依赖的方式抑制。然而，线粒体缺失的细胞仍然能够对缺氧作出反应，上调RBM3和CIRP。因此，RBM3和CIRP通过一种既不涉及HIF-1也不涉及线粒体的机制对缺氧作出适应性表达。
