Facilitation of a structural transition in the polypurine/polypyrimidine tract within the proximal promoter region of the human VEGF gene by the presence of potassium and G-quadruplex-interactive agents.
Facilitation of a structural transition in the polypurine/polypyrimidine tract within the proximal promoter region of the human VEGF gene by the presence of potassium and G-quadruplex-interactive agents.
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DOI：
10.1093/nar/gki917
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发表时间：
2005
影响因子：
14.9
通讯作者：
Hurley LH
中科院分区：
文献类型：
--
作者：
Sun D;Guo K;Rusche JJ;Hurley LH
人血管内皮生长因子（VEGF）基因的近端启动子区含有一个多嘌呤/多嘧啶区，可作为Sp1和Egr-1转录因子的多个结合位点。这道含有一个富含鸟嘌呤的序列，由四个连续的三个或更多个连续的鸟嘌呤组成，由一个或多个碱基分开，对应于形成分子内G-四链体的一般基序。在这项研究中，我们观察到的寡聚体双链体DNA含有这个区域的逐步解旋成单链形式的存在下，氯化钾和G-四链体相互作用剂TMPyP 4和端粒，这表明动态性质的条件下，有利于G-四链体结构的形成，这道。随后的足迹研究与DNA酶I和S1核酸酶使用超螺旋质粒DNA含有人VEGF启动子区也揭示了一个长的保护区，包括鸟嘌呤丰富的序列，在KCl和端粒。值得注意的是，在KCl和端粒酶抑制素存在下，在预测的G-四链体形成区的3′-侧残基处观察到对两种核酸酶的显著超敏反应，这表明富含鸟嘌呤序列周围的人VEGF近端启动子区的构象发生了改变。相比之下，当特定的点突变被引入到特定的鸟嘌呤残基内的G-四链体形成区域（Sp1结合位点），以消除G-四链体形成能力，两个核酸酶对突变的人VEGF近端启动子区域的反应性几乎是相同的，即使在端粒抑素与KCl的存在下。野生型和突变体序列的这种比较强烈表明，高度组织化的二级结构，如G-四链体的人VEGF启动子区的富G区域内的形成是负责观察到的人VEGF基因的多嘌呤/多嘧啶道内的两种核酸酶的反应性的变化。CD实验进一步证实了人VEGF启动子中该富含G的序列形成G-四链体结构。总的来说，我们的研究结果提供了强有力的证据表明，特定的G-四链体结构可以自然地形成的人VEGF启动子区域的多嘌呤/多嘧啶道内的G-丰富的序列，提高了VEGF基因的转录控制可以通过G-四链体相互作用剂调制的可能性。
The proximal promoter region of the human vascular endothelial growth factor (VEGF) gene contains a polypurine/polypyrimidine tract that serves as a multiple binding site for Sp1 and Egr-1 transcription factors. This tract contains a guanine-rich sequence consisting of four runs of three or more contiguous guanines separated by one or more bases, corresponding to a general motif for the formation of an intramolecular G-quadruplex. In this study, we observed the progressive unwinding of the oligomer duplex DNA containing this region into single-stranded forms in the presence of KCl and the G-quadruplex-interactive agents TMPyP4 and telomestatin, suggesting the dynamic nature of this tract under conditions which favor the formation of the G-quadruplex structures. Subsequent footprinting studies with DNase I and S1 nucleases using a supercoiled plasmid DNA containing the human VEGF promoter region also revealed a long protected region, including the guanine-rich sequences, in the presence of KCl and telomestatin. Significantly, a striking hypersensitivity to both nucleases was observed at the 3′-side residue of the predicted G-quadruplex-forming region in the presence of KCl and telomestatin, indicating altered conformation of the human VEGF proximal promoter region surrounding the guanine-rich sequence. In contrast, when specific point mutations were introduced into specific guanine residues within the G-quadruplex-forming region (Sp1 binding sites) to abolish G-quadruplex-forming ability, the reactivity of both nucleases toward the mutated human VEGF proximal promoter region was almost identical, even in the presence of telomestatin with KCl. This comparison of wild-type and mutant sequences strongly suggests that the formation of highly organized secondary structures such as G-quadruplexes within the G-rich region of the human VEGF promoter region is responsible for observed changes in the reactivity of both nucleases within the polypurine/polypyrimidine tract of the human VEGF gene. The formation of the G-quadruplex structures from this G-rich sequence in the human VEGF promoter is further confirmed by the CD experiments. Collectively, our results provide strong evidence that specific G-quadruplex structures can naturally be formed by the G-rich sequence within the polypurine/polypyrimidine tract of the human VEGF promoter region, raising the possibility that the transcriptional control of the VEGF gene can be modulated by G-quadruplex-interactive agents.
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影响因子：
15
作者：
Han, HY;Langley, DR;Hurley, LH
通讯作者：
Hurley, LH
影响因子：
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作者：
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通讯作者：
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影响因子：
11.4
作者：
HOLSTEGE, FCP;TANTIN, D;TIMMERS, HTM
通讯作者：
TIMMERS, HTM
影响因子：
4.8
作者：
Pal, S;Datta, K;Mukhopadhyay, D
通讯作者：
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影响因子：
14.9
作者：
LAFYATIS, R;DENHEZ, F;ROBERTS, A
通讯作者：
ROBERTS, A