SIRT6调节GATA4转录活性在心脏衰老及心力衰竭中的作用研究

Older people suffer from heart failure predominantly triggered by cardiac aging. Deacetylase SIRT6 is cardiac-protective. Our preliminary data showed that SIRT6 could interact with GATA4 and significantly increased acetylation level of GATA4. GATA4 is a transcription factor with a key role in heart development and regeneration. Its function in cardiac aging is unclear. This project is planning to investigate the molecular mechanism of SIRT6-mediated GATA4 acetylation, and to screen its specific lysine site for SIRT6 regulation. Our further study is to explore the effect of SIRT6-mediated GATA4 acetylation on GATA4 transcriptional activity by performing electrophoretic mobility shift assay, chromatin immunoprecipitation and luciferase reporter assay. Since GATA4 was significantly down-regulated in aged and SIRT6-depleted heart as shown in our preliminary result, we will assess the role of GATA4 in cardiac aging and doxorubicin-induced heart failure by utilizing gene knockout and transgenic mouse models. By comparing gene expression profile in heart tissue from SIRT6-/- and SIRT6-/-; Gata4 Myh-Tg mice, we will address how SIRT6 depletion induces cardiac hypotrophy and GATA4 rescues premature aging phenotype in SIRT6-/- mice. Collectively，these study will uncover a novel pathway regulating cardiac aging, providing new prospects for the drug development against aging-related heart disease.

老年人心脏老化易发心衰。去乙酰化酶SIRT6具心血管保护功能。本课题前期研究发现SIRT6可结合并增强GATA4乙酰化。GATA4是心脏发育和心肌再生的重要转录因子，与心脏衰老的关联性并不清楚。本课题拟研究SIRT6增强GATA4乙酰化的分子机制；筛选受调控的乙酰化位点；通过DNA凝胶迁移（EMSA）、ChIP和荧光素酶报告系统检测SIRT6介导GATA4乙酰化对其转录活性的影响。预实验发现GATA4在老年和SIRT6-/-小鼠心脏中蛋白含量显著降低。据此本研究将采用基因缺失与转基因小鼠模型，进而探讨GATA4在心脏衰老及阿霉素诱导心衰中的作用；通过转录组学比较SIRT6-/-和SIRT6-/-;GATA4Myh-Tg小鼠心脏的基因表达谱，探讨SIRT6缺失诱发心肌肥大和GATA4回补早衰表型的机制。本项目预期成果将揭示心脏衰老分子机制的新通路，对研发老年性心脏疾病的靶向药物提供新思路。

化疗药物阿霉素（DOX）是临床治疗肿瘤常用药物，由于强的心脏毒性限制了其使用范围，而老年人尤为敏感。去乙酰化酶SIRT6作为长寿基因，具有保护心脏功能。本项目旨在探究SIRT6抵抗DOX诱导的心衰中的作用机制。在本项目的资助下，我们围绕“去乙酰化酶SIRT6和心肌重要转录因子GATA4对心脏保护功能的调控机制”展开研究，取得了多项成果。.利用SIRT6心脏特异性缺失小鼠模型和体外原代心肌细胞实验，我们发现SIRT6作为DOX诱导心衰的早期响应分子，通过心脏重要转录因子GATA4调控抗凋亡基因的表达。过表达SIRT6小鼠心脏则可以显著抑制由DOX诱导的心肌细胞抗凋亡基因的下调。分子生化实验显示SIRT6与GATA4互作，但却显著增强了GATA4的乙酰化。我们进一步发现TIP60是参与SIRT6介导的GATA4乙酰化的乙酰转移酶，DOX处理破坏了三者相互作用，从而降低GATA4调控的抗凋亡基因表达。反之，GATA4蛋白结合SIRT6抑制了SIRT6去乙酰化酶活性，GATA4与SIRT6在抗凋亡启动子区域的结合不影响染色质的可及性。.因此，本项目研究揭示了SIRT6不依赖酶活性的心脏保护功能；阐述了SIRT6介导转录因子GATA4对于抗凋亡基因正向表达调控的分子机制；发现了新型的GATA4活性调控的乙酰转移酶TIP60；阐明了DOX破坏TIP60-SIRT6-GATA4复合体是引起心肌毒性的潜在原因。这些研究成果进一步丰富了SIRT6作为长寿基因的生理功能基础，也为临床辅助放化疗药物开发提供新思路。