Cardiac TranscriptionFactor Interactomes regulating cardiac development&Reprogram

信号、转录和转录后事件调节心脏细胞在早期心脏发生过程中的命运决定。这类事件的中断可能会导致先天性心脏畸形。特别是，人类转录因子的突变，如GATA4、TBX5、NKX2-5和NOTCHI，会导致儿童心脏病。在成年心脏中，胚胎通路在应激条件下被重新激活，GATA4和MEF2C在心肌肥厚时的转录反应中发挥核心作用。最近的研究强调了蛋白质-蛋白质相互作用(PPI)在决定DNA结合转录因子的转录输出中的重要性。然而，滴定心脏转录因子效应的复杂的PPI还没有得到系统的探索。在该PPG的前一次资助期间，我们的发现主要集中在心脏发育过程中Notch和Wnt信号通路成员之间的PPI。此外，我们报道了包括Gata4、MEF2C和Tbx5在内的心脏发育转录因子的组合可以在成人中将非肌肉细胞重新编程为新的心肌细胞样细胞，从而导致损伤后的心脏再生；Hand2的添加进一步改善了重新编程。PPG更新应用的项目1将测试Gata4、MEF2C、Tbx5和Hand2相互作用复杂的假设，以及在心脏分化和心脏重编程过程中调节转录输出的其他关键因素。具体的措施是：(1)绘制小鼠胚胎干细胞向心肌细胞分化过程中涉及Gata4、MEF2C、Tbx5和Hand2的交互作用组图；(2)确定相互作用对Gata4、Tbx5和Hand2的中心依赖性，以及GATA4致病错义突变对相互作用组的影响；以及(3)确定涉及Gata4、MEF2C、Tbx5和Hand2的PPI对特定基因组位置的功能影响，并将PPI与心脏分化和重新编程期间转录因子的基因组占有率整合。该项目将揭示心脏基因调控的潜在机制，并将提供潜在的干预点，以积极或消极的转录活性滴定。

Signaling, transcriptional, and post-transcriptional events regulate cardiac cell fate decisions during early cardiogenesis. Disruption of such events can lead to congenital heart malformations. In particular, human mutations in transcription factors, such as GATA4, TBX5, NKX2-5 and NOTCHI, result in heart disease in children. Embryonic pathways are reactivated under stress in adult hearts, with GATA4 and MEF2C playing central roles in the transcripfional response during cardiac hypertrophy. Recent studies highlight the importance of protein-protein interactions (PPI) in dictating the transcripfional output of DNA binding transcription factors. However, the complex PPIs that titrate effects of cardiac transcription factors have not been systematically explored. During the previous funding period of this PPG, our discoveries focused on PPIs between members ofthe Notch and Wnt signaling pathways during cardiac development. In addition, we reported that a combination of cardiac developmental transcription factors, including Gata4, Mef2c and Tbx5, could reprogram non-muscle cells into new cardiomyocyte-like cells in the adult, resulting in cardiac regeneration after injury; the addition of Hand2 further improves reprogramming. Project 1 of this PPG renewal application will test the hypothesis that Gata4, Mef2c, Tbx5, and Hand2 have complex interactions with one another and other key factors to regulate the transcripfional output during cardiac differentiation and cardiac reprogramming. The specific alms are (1) to develop a comprehensive map ofthe Interactome involving Gata4, Mef2c, Tbx5 and Hand2 during cardiac differentiation of mouse embryonic stem (ES) cells into cardiomyocytes; (2) determine the central dependency of interactomes on Gata4, Tbx5 and Hand2, and the consequences of disease-causing missense mutations In GATA4 on the interactome; and (3) determine the functional consequences of PPIs involving Gata4, Mef2c, Tbx5 and Hand2 on specific genomic loci and integrate PPIs with genome occupancy ofthe transcription factors during cardiac differentiation and reprogramming. This project will reveal mechanisms underlying cardiac gene regulation and will provide potential points of intervention to positively or negatively titrate transcriptional activity.