Inhibition of hypoxia-inducible factor (HIF) hydroxylases by citric acid cycle intermediates -: Possible links between cell metabolism and stabilization of HIF
Inhibition of hypoxia-inducible factor (HIF) hydroxylases by citric acid cycle intermediates -: Possible links between cell metabolism and stabilization of HIF
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DOI：
10.1074/jbc.m610415200
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发表时间：
2007-02-16
影响因子：
4.8
通讯作者：
Myllyharju, Johanna
中科院分区：
文献类型：
--
作者：
Koivunen, Peppi;Hirsila, Maija;Myllyharju, Johanna
低氧诱导因子(HIF)的稳定性和转录活性受3个HIF-P4Hs和1个HIF天冬酰胺基羟基酶(FIH)催化的两个氧依赖事件的调节。我们通过分析柠檬酸循环中间体抑制纯化的人HIF-P4Hs和FIH的能力，研究了代谢途径和HIF羟基酶之间的可能联系。富马酸和琥珀酸是三种HIF-P4H的体外抑制剂，富马酸的K-I值在50-80 mU M之间，琥珀酸的K-I值在350-460 mU M之间，但都不能抑制FIH。草酰乙酸酯是三种HIF-P4H的附加抑制剂，K-I值为400-1000 mU M，柠檬酸盐为HIF-P4H-3的柠檬酸盐，柠檬酸盐是FIH最有效的抑制剂，K-I为110 mU M。在富马酸水合酶(FH)缺乏症患者和使用小干扰RNA沉默FH后，培养的成纤维细胞中也发现了类似的变化，尽管变化要小得多。在用琥珀酸二乙酯或二甲酯培养细胞时，没有观察到这样的影响。由于FIH不被富马酸抑制，我们的数据表明，即使阻止辅活化子p300的结合，HIF的转录活性也相当高。我们的数据也支持最近的观点，即在FH和琥珀酸脱氢酶缺陷的肿瘤中，富马酸和琥珀酸水平的增加分别可以抑制HIF-P4Hs，从而稳定HIF-αS，并对肿瘤病理产生影响。
The stability and transcriptional activity of the hypoxia-inducible factors (HIFs) are regulated by two oxygen-dependent events that are catalyzed by three HIF prolyl 4-hydroxylases (HIF-P4Hs) and one HIF asparaginyl hydroxylase (FIH). We have studied possible links between metabolic pathways and HIF hydroxylases by analyzing the abilities of citric acid cycle intermediates to inhibit purified human HIF-P4Hs and FIH. Fumarate and succinate were identified as in vitro inhibitors of all three HIF-P4Hs, fumarate having K-i values of 50-80 mu M and succinate 350-460 mu M, whereas neither inhibited FIH. Oxaloacetate was an additional inhibitor of all three HIF-P4Hs with K-i values of 400-1000 mu M and citrate of HIF-P4H-3, citrate being the most effective inhibitor of FIH with a K-i of 110 mu M. Culturing of cells with fumarate diethyl or dimethyl ester, or a high concentration of monoethyl ester, stabilized HIF-1 alpha and increased production of vascular endothelial growth factor and erythropoietin. Similar, although much smaller, changes were found in cultured fibroblasts from a patient with fumarate hydratase (FH) deficiency and upon silencing FH using small interfering RNA. No such effects were seen upon culturing of cells with succinate diethyl or dimethyl ester. As FIH was not inhibited by fumarate, our data indicate that the transcriptional activity of HIF is quite high even when binding of the coactivator p300 is prevented. Our data also support recent suggestions that the increased fumarate and succinate levels present in the FH and succinate dehydrogenase-deficient tumors, respectively, can inhibit the HIF-P4Hs with consequent stabilization of HIF-alpha s and effects on tumor pathology.