Transcriptional regulation of cardiac conduction system morphogenesis

项目摘要/摘要 电脉冲的传播，它协调有节奏的和同步的心脏收缩 促进体循环是由一种特殊的结构和细胞类型调节的，心脏传导 系统(CCS)，并以时空精确的方式工作心肌细胞。先天性颈椎畸形 CCS和CCS动态平衡失调可导致CCS功能障碍，导致生命危险 心律失常和增加儿童和成人的死亡风险。全基因组关联研究 在人类各种心律失常患者中均发现异常心律失常密切相关。 心电和许多离子通道、缝隙连接蛋白、肌肉结构蛋白以及一些关键的 转录因子在心脏发育中的作用及其特异性、分化和稳态 CCS的维持，其中包括碱性螺旋-环-螺旋(BHLH)转录因子Hand1 和T-box转录因子Tbx20。Hand1在发育早期的心脏中表达，是必需的 用于心脏形态发生。Tbx20被证明与多种心脏事件和心脏疾病密切相关。 功能。有趣的是，尽管Hand1和Tbx20以前没有与CCS开发中的角色相关联 或功能，Gwas分析揭示了HAND1和TBX20内的单核苷酸多态(SNPs) 与QRS持续时间延长和强连锁不平衡有关。适用于HAND1和TBX20 这些SNP是基因间的，提示在这些基因的转录调控中发挥作用。事实上，我们有 发现了一个带有GATA4和T-box结合位点的左心室(LV)Hand1增强子，它位于 位于已报道的SNP之间的保守非编码序列(CNS)。突变的小鼠有 含有这种增强子的基因编辑缺失会导致QRS延长。支持这一发现，我们有 最近观察到Hand1在出生后心脏的CCS结构中特异表达。 Tbx20的转录调控在多个心肌细胞系中表达。心肌得或失- 功能研究显示QRS时限改变和严重心律失常。总而言之，我们压倒一切的 假设Hand1和Tbx20协调并维持心脏的空间和时间控制 传导系统的发育和功能通过平行或单基因调控途径实现。我们会 用以下具体目标测试这一假设：1)测试成人Hand1表达的假设 在CCS内需要维持正常的QRS持续时间；2)测试Tbx20的假设 在CCS发育和动态平衡以及维持导电性方面的作用 心肌细胞的功能；3)检测Hand1和Tbx20是否作为关键的转录调节因子 CCS的发育和CCS的生理功能有共同或不同的调节途径。

PROJECT SUMMARY/ABSTRACT The propagation of electrical impulses that coordinate the rhythmic and synchronized cardiac contractions to facilitate systemic circulation is regulated by a specialized structures and cell types, the cardiac conduction system (CCS), and working cardiomyocytes in a spatial-temporally precise manner. Congenital defects of the CCS and dysregulation of CCS homeostasis can lead to CCS dysfunction, causing life threatening arrhythmias and increasing the risk of death in both children and adults. Genome Wide Association studies (GWAS) in human patients with various arrhythmias have revealed a close association between abnormal ECG and many ion channels, gap junction proteins, muscle structural proteins as well as a number of critical transcription factors that function in cardiac development and the specification, differentiation and homeostatic maintenance of the CCS, among them include the basic Helix-loop-Helix (bHLH) transcription factor Hand1 and the T-box transcription factor Tbx20. Hand1 is expressed in the early developing hearts and is essential for cardiac morphogenesis. Tbx20 is shown to be critically involved in multiple cardiogenic events and cardiac function. Interestingly, despite Hand1 and Tbx20 are not previously associated with roles in CCS development or function, GWAS analysis revealed single nucleotide polymorphisms (SNPs) within both HAND1 and TBX20 associated with prolonged QRS duration with strong linkage disequilibrium. For both HAND1 and TBX20 these SNPs are intergenic suggesting roles in transcriptional regulation of these genes. Indeed, we have identified a left ventricle (LV) Hand1 enhancer with GATA4 and T-box binding sites that resides within a conserved non-coding sequence (CNS) that lies in between the reported SNPs. Mutant mice that have harbored a gene-edited deletion of this enhancer result in a prolonged QRS. Supporting this finding, we have recently observed that Hand1 is specifically expressed within CCS structures in postnatal hearts. Transcriptional regulation of Tbx20 is expressed in multiple cardiac cell lineages. Myocardial gain- or loss-of- function studies have shown altered QRS duration and severe arrhythmia. Collectively, our overriding hypothesis is that Hand1 and Tbx20 coordinate and maintain the spatial and temporal control of the cardiac conductive system development and function via either parallel or single gene regulatory pathways. We will test this hypothesis with the following specific Aims: 1) to test the hypothesis that adult expression of Hand1 within the CCS is required for maintaining a normal QRS duration; 2) to test the hypothesis that Tbx20 functions in the development and homeostasis of the CCS as well as the maintenance of the conductive function of cardiomyocytes; 3) to test whether Hand1 and Tbx20 serves as key transcriptional regulators for CCS development and CCS physiological function in a common or different regulatory pathways.