Hypoxia-induced oxidative base modifications in the VEGF hypoxia-response element are associated with transcriptionally active nucleosomes.
Hypoxia-induced oxidative base modifications in the VEGF hypoxia-response element are associated with transcriptionally active nucleosomes.
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DOI：
10.1016/j.freeradbiomed.2008.09.038
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发表时间：
2009-02-01
影响因子：
7.4
通讯作者：
Gillespie, Mark N.
中科院分区：
文献类型：
--
作者：
Ruchko, Mykhaylo V.;Gorodnya, Olena M.;Pastukh, Viktor M.;Swiger, Brad M.;Middleton, Natavia S.;Wilson, Glenn L.;Gillespie, Mark N.
关键词：
在缺氧肺动脉内皮细胞中产生的活性氧物质（ROS）引起VEGF基因的缺氧反应元件（HRE）中的瞬时氧化碱基修饰，其与VEGF mRNA表达的诱导具有显著的关系（FASEB J 19：387-394，2005）。如果这种碱基修饰确实与转录调控有关，那么它们应该在与转录活性核小体相关的HRE序列中检测到。与微球菌核酸酶消化制备的核小体组分相关的VEGF HRE的Southern印迹分析表明，缺氧将一些HRE序列从多核小体重新分配到转录活性的单核小体和双核小体组分。一个简单的PCR方法显示，VEGF HRE序列窝藏氧化碱基修饰被发现专门在单核小体。低氧诱导的活性氧生成的抑制与myxathiozol防止形成的氧化碱基修饰，但不是重新分配的HRE序列到单和双核小体部分。组蛋白去乙酰化酶抑制剂，抑制素A，引起HRE序列的保留在紧凑的核小体部分，并防止形成氧化碱基修饰。这些发现表明，缺氧诱导的氧化应激针对VEGF HRE需要的序列被重新定位成单核小体，并支持前景，该序列中的氧化修饰是转录激活的重要步骤。
Reactive oxygen species (ROS) generated in hypoxic pulmonary artery endothelial cells cause transient oxidative base modifications in the hypoxic response element (HRE) of the VEGF gene that bear a conspicuous relationship to induction of VEGF mRNA expression (FASEB J 19: 387-394, 2005). If such base modifications are indeed linked to transcriptional regulation, then they should be detected in HRE sequences associated with transcriptionally active nucleosomes. Southern blot analysis of the VEGF HRE associated with nucleosome fractions prepared by micrococcal nuclease digestion indicated that hypoxia redistributed some HRE sequences from multi-nucleosomes to transcriptionally-active mono- and di-nucleosome fractions. A simple PCR method revealed that VEGF HRE sequences harboring oxidative base modifications were found exclusively in mono-nucleosomes. Inhibition of hypoxia-induced ROS generation with myxathiozol prevented formation of oxidative base modifications but not the redistribution of HRE sequences into mono- and di-nucleosome fractions. The histone deacetylase inhibitor, trichostatin A, caused retention of HRE sequences in compacted nucleosome fractions and prevented formation of oxidative base modifications. These findings suggest that the hypoxia-induced oxidant stress directed at the VEGF HRE requires the sequence to be repositioned into mono-nucleosomes, and support the prospect that oxidative modifications in this sequence are an important step in transcriptional activation.
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