The i-motif in the bcl-2 P1 promoter forms an unexpectedly stable structure with a unique 8:5:7 loop folding pattern.
The i-motif in the bcl-2 P1 promoter forms an unexpectedly stable structure with a unique 8:5:7 loop folding pattern.
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DOI：
10.1021/ja9076292
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发表时间：
2009-12-09
影响因子：
15
通讯作者：
Hurley, Laurence H.
中科院分区：
文献类型：
--
作者：
Kendrick, Samantha;Akiyama, Yoshitsugu;Hecht, Sidney M.;Hurley, Laurence H.
bcl-2原癌基因的转录调控是高度复杂的，大部分转录由P1启动子位点和多个调控蛋白的相互作用驱动。P1位点上游富含鸟嘌呤和胞嘧啶（GC-rich）的区域已被证明是bcl-2启动子活性的组成部分，因为该区域的缺失或突变会显著增加转录。这种富含gc的元件由六个连续的鸟嘌呤和胞嘧啶组成，它们分别具有采用DNA二级结构g -四重体和i-基序的潜力。我们的实验室先前已经证明，bcl-2启动子的富含多嘌呤的链可以形成三种不同g -四重结构的混合物。在目前的研究中，我们证明了互补的富含多聚嘧啶的链能够形成一个主要的分子内i-motif DNA二级结构，过渡pH为6.6。利用突变研究、圆二色性光谱和热稳定性分析对i-motif折叠模式进行表征，发现在pH为6.1时，其主要结构为8:5:7环。溴的化学足迹进一步支持了折叠模式。此外，一项涉及在每个胸腺嘧啶位置依次加入荧光胸腺嘧啶类似物的新实验提供了i-motif的顶部环区域内的盖层结构的证据。bcl-2启动子区域富含gc元素形成DNA二级结构的潜力表明，从B-DNA到非B-DNA构象的转变可能在bcl-2转录调控中发挥重要作用。此外，i-motif结构中两个相邻的大横向环为蛋白质和小分子识别提供了意想不到的机会。
Transcriptional regulation of the bcl-2 proto-oncogene is highly complex, with the majority of transcription driven by the P1 promoter site and the interaction of multiple regulatory proteins. A guanine- and cytosine-rich (GC-rich) region directly upstream of the P1 site has been shown to be integral to bcl-2 promoter activity, as deletion or mutation of this region significantly increases transcription. This GC-rich element consists of six contiguous runs of guanines and cytosines that have the potential to adopt DNA secondary structures, the G-quadruplex and i-motif, respectively. Our laboratory has previously demonstrated that the polypurine-rich strand of the bcl-2 promoter can form a mixture of three different G-quadruplex structures. In this current study, we demonstrate that the complementary polypyrimidine-rich strand is capable of forming one major intramolecular i-motif DNA secondary structure with a transition pH of 6.6. Characterization of the i-motif folding pattern using mutational studies coupled with circular dichroic spectra and thermal stability analyses revealed an 8:5:7 loop conformation as the predominant structure at pH 6.1. The folding pattern was further supported by chemical footprinting with bromine. In addition, a novel assay involving the sequential incorporation of a fluorescent thymine analog at each thymine position provided evidence of a capping structure within the top loop region of the i-motif. The potential of the GC-rich element within the bcl-2 promoter region to form DNA secondary structures suggests that the transition from the B-DNA to non-B-DNA conformation may play an important role in bcl-2 transcriptional regulation. Furthermore, the two adjacent large lateral loops in the i-motif structure provide an unexpected opportunity for protein and small molecule recognition.
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DOI：
10.1039/jr9480001759
发表时间：
1948-01-01
期刊：
JOURNAL OF THE CHEMICAL SOCIETY
影响因子：
--
作者：
CURD, FHS;LANDQUIST, JK;ROSE, FL
通讯作者：
ROSE, FL
影响因子：
14.9
作者：
GROEBE, DR;UHLENBECK, OC
通讯作者：
UHLENBECK, OC
影响因子：
14.9
作者：
Guo K;Gokhale V;Hurley LH;Sun D
通讯作者：
Sun D
影响因子：
2.9
作者：
HAASNOOT, CAG;DEBRUIN, SH;VANBOOM, JH
通讯作者：
VANBOOM, JH
影响因子：
2.2
作者：
Cashman, Derek J.;Buscaglia, Robert;Lewis, Edwin A.
通讯作者：
Lewis, Edwin A.