G-四联体的形成和稳定在松弛素的表达调控中的作用及机制

Myocardial fibrosis is a pathological basis for a variety of heart diseases. Previous studies have shown that relaxin prevents and reverses cardiac fibrosis. Endogenous expression of relaxin was elevated in the setting of heart disease, the extent of such upregulation, however, is insufficient to exert compensatory actions and the mechanism regulating relaxin expression is poorly defined. It has been demonstrated by a previous study that STAT3 negatively regulates relaxin expression. We recently found in the relaxin gene promoter region presence of repeated guanine (G)-rich sequences, which allowed formation and stabilization of G-quadruplexes with the addition of a G-quadruplex interactive ligand. The G-rich sequences of relaxin gene were next to the STAT3 binding site separated by 11 nucleotides. Thus, we hypothesize that the formation and stabilization of the G-quadruplexe by the ligand can influence the binding of STAT3 with relaxin gene, increase relaxin expression and achieve anti-fibrotic effect. In the present study, we aims at elucidating effect of formation and stabilization of G-quadruplexes on the upregulation of endogenous relaxin expression by ChIP assay and CD spectrum in vitro and in vivo.

心肌纤维化是多种心血管疾病的重要病理基础。大量研究表明外源性松弛素可以显著抑制心肌纤维化，并且在疾病动物模型中，升高的内源性松弛素不足以阻止心肌纤维化的进展。因此我们提出能否促进内源性松弛素的表达进而抑制心肌纤维化的发展？目前关于上调内源性松弛素表达的机制尚不清楚。已有研究证实STAT3负性调控松弛素的表达。我们近期研究发现在松弛素基因的启动子区STAT3结合位点下游11个碱基处存在一段富G的重复序列，该序列可以在特定的配基作用下可以形成特殊的DNA二级结构-G四联体。由此，我们大胆假设通过特定配基诱导形成并稳定G-四联体结构可以干扰STAT3与松弛素启动子区的结合，从而上调松弛素的表达，发挥抗心肌纤维化作用。本研究拟通过ChIP、圆二色谱等实验方法，在体外、细胞和整体水平阐明G-四联体的形成和稳定上调松内源性松弛素基因的表达水平。

本课题研究发现心脏重塑相关的重要基因（松弛素，RLN）可以被G-四链体的形成和稳定所调控。研究成果发表于Endocrinology 2012上，并受邀在2013年意大利的《松弛素论坛》做了“G-四链体与心脏重塑”的专题报告。在上述发现的基础上，结合申请者近期的发现 “miR-27b可以调控松弛素受体RXP1的表达水平”，提出了基于microRNA的松弛素受体调控新机制，并获得国家自然科学基金重大项目预探索项目的资助。