Instituto de Biología Molecular y Celular de Rosario (IBR), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET)-Area Biología General, Dpto. de Ciencias Biológicas, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Suipacha 531, S2002LRK Rosario, Argentina.
Biochem J. 2010 May 27;428(3):491-8. doi: 10.1042/BJ20100038.
G-rich sequences that contain stretches of tandem guanines can form four-stranded, intramolecular stable DNA structures called G-quadruplexes (termed G4s). Regulation of the equilibrium between single-stranded and G4 DNA in promoter regions is essential for control of gene expression in the cell. G4s are highly stable structures; however, their folding kinetics are slow under physiological conditions. CNBP (cellular nucleic-acid-binding protein) is a nucleic acid chaperone that binds the G4-forming G-rich sequence located within the NHE (nuclease hypersensitivity element) III of the c-Myc proto-oncogene promoter. Several reports have demonstrated that CNBP enhances the transcription of c-Myc in vitro and in vivo; however, none of these reports have assessed the molecular mechanisms responsible for this control. In the present study, by means of Taq polymerase stop assays, electrophoretic mobility-shift assays and CD spectroscopy, we show that CNBP promotes the formation of parallel G4s to the detriment of anti-parallel G4s, and its nucleic acid chaperone activity is required for this effect. These findings are the first to implicate CNBP as a G4-folding modulator and, furthermore, assign CNBP a novel mode-of-action during c-Myc transcriptional regulation.
富含鸟嘌呤的序列包含串联鸟嘌呤，可以形成称为 G-四链体 (G4s) 的四链、分子内稳定的 DNA 结构。在启动子区域中，调节单链和 G4 DNA 之间的平衡对于控制细胞中的基因表达至关重要。G4 是高度稳定的结构；然而，它们在生理条件下的折叠动力学较慢。CNBP（细胞核酸结合蛋白）是一种核酸伴侣，可结合 c-Myc 原癌基因启动子中 NHE（核酸酶超敏元件）III 内的 G4 形成富含鸟嘌呤的序列。有几项报道表明，CNBP 增强了体外和体内 c-Myc 的转录；然而，这些报道都没有评估负责这种控制的分子机制。在本研究中，通过 Taq 聚合酶停止测定、电泳迁移率变动分析和 CD 光谱分析，我们表明 CNBP 促进平行 G4 的形成，而不利于反平行 G4 的形成，其核酸伴侣活性是产生这种效果所必需的。这些发现首次将 CNBP 作为 G4 折叠调节剂，并进一步在 c-Myc 转录调控过程中赋予 CNBP 一种新的作用模式。
