联合CRISPR-Cas9及AAV9技术研究YAP/TAZ应力应答途径在心脏病理性重构中的调控机制

Cardiac pathological remodeling participates in the procedure of heart failure as an important role. However, no specific target treatment protocol has been initiated. During the remodeling process, cardiac phenotypic genes demonstrate a “Return to Fetal Gene Program”, which results in irreversible pathological status. Our preliminary research, based on a pioneer fetal heart failure model, revealed an antithetical transformation of cardiac phenotypic genes as “Activated Adult Gene Program” under responding to fetal low level of mechanical stress. Besides, positive regression claimed adult phenotypes persisted advantageous heart function. Moreover, the core factors YAP/TAZ in responding mechanical stress have also been proved diverse expressions and involved in the disrupting compensation between to different stresses. Besides, with the dramatic progress in gene editing technology, based on CRISPR-Cas and AAV9, it would realize the rapid, accurate and efficient gene editing in vivo. In this project, we suppose to establish the model of YAP/TAZ mechanical responding pathway in regulating of fetal and adult phenotypic gene program during cardiac pathological remodeling in collaboration with CRISPR-Cas9 and AAV9. From the insight of emerging mechanotransduction pathway, we will get entire understanding of YAP/TAZ in shifting fetal and adult phenotypic gene program in responding to abnormal hemodynamics, and further to identify the potential therapeutic targets to switch the expression of gene program intelligently. Our results would elucidate an originative sight in reviewing current opinion on heart failure and propel its treatment forward.

心脏病理性重构是心力衰竭疾病进程中的重要环节，目前仍缺乏特异性干预手段。该进程中心脏表型基因发生 “重返胎儿基因程序”，并导致不可逆病理性重构。我们前期研究显示，胎儿期低应力负荷心力衰竭模型中心脏表型基因表达与成年期具有差异性，呈现为“提前激活成人基因程序”，并与心功能维持呈正相关；并发现细胞应力应答途径关键蛋白YAP/TAZ亦参与其中，且同样出现差异性表达。同时基于基因编辑手段的快速进展，联合CRISPR-Cas9及AAV9技术可以于在体水平实现快速、准确且高效的基因干预。本项目拟通过应用CRISPR-Cas9及AAV9技术，阐明YAP/TAZ介导应力应答途径在心脏病理性重构中对胎儿/成年基因程序作用模式，从力学信号应答途径入手，阐明病理状态下YAP/TAZ应力应答途径在异常心脏收缩表型基因表达中的作用，筛选有效开关胎儿/成人基因程序特异性靶点，以全新视角推进心力衰竭的治疗。

异常应力负荷可致线粒体结构与功能障碍从而诱发心脏病理性重构，但其应力转导途径及分子机制仍不清楚。本项目研究发现异常应力负荷可显著激活心肌细胞YAP，并与VGLL4竞争性结合TEAD1；形成YAP-TEAD1复合物，结合于Drp1、Mfn1基因启动子并上调其转录活性，驱动线粒体动态异常而导致心脏病理性重构。因此本研究以力学信号转导途径对线粒体动态的调控机制为切入点，利用ATAC-seq、bioChIP-seq及多种Cre-Loxp基因小鼠等实验方法，阐明异常应力负荷下VGLL4/YAP表达及线粒体结构与功能；明确了VGLL4/YAP-TEAD1可调控Tfam、Mfn1、Mfn2基因活性以驱动线粒体动态失衡是异常应力负荷致心脏病理性重构的核心分子机制；初步探究干预VGLL4/YAP以稳定线粒体动态平衡从而达到治疗心脏病理性重构的可行性，从全新视角推进心肌肥厚的治疗。