TBX5 drives Scn5a expression to regulate cardiac conduction system function

Abstract

Cardiac conduction system (CCS) disease, which results in disrupted conduction and impaired cardiac rhythm, is common with significant morbidity and mortality. Current treatment options are limited, and rational efforts to develop cell-based and regenerative therapies require knowledge of the molecular networks that establish and maintain CCS function. Recent genome-wide association studies (GWAS) have identified numerous loci associated with adult human CCS function, including TBX5 and SCN5A. We hypothesized that TBX5, a critical developmental transcription factor, regulates transcriptional networks required for mature CCS function. We found that deletion of Tbx5 from the mature murine ventricular conduction system (VCS), including the AV bundle and bundle branches, resulted in severe VCS functional consequences, including loss of fast conduction, arrhythmias, and sudden death. Ventricular contractile function and the VCS fate map remained unchanged in VCS-specific Tbx5 knockouts. However, key mediators of fast conduction, including Na v 1.5, which is encoded by Scn5a, and connexin 40 (Cx40), demonstrated Tbx5-dependent expression in the VCS. We identified a TBX5-responsive enhancer downstream of Scn5a sufficient to drive VCS expression in vivo, dependent on canonical T-box binding sites. Our results establish a direct molecular link between Tbx5 and Scn5a and elucidate a hierarchy between human GWAS loci that affects function of the mature VCS, establishing a paradigm for understanding the molecular pathology of CCS disease. IntroductionThe cardiac conduction system (CCS) consists of a network of specialized cardiomyocytes that generate and propagate the electrical impulses that organize cardiac contraction. The CCS is composed of the slowly propagating atrial nodes, including the sinoatrial (SA) and atrioventricular (AV) nodes, and the rapidly propagating ventricular conduction system (VCS), including the AV (His) bundle and right and left bundle branches. The VCS is uniquely adapted for fast conduction in order to rapidly transmit the electrical impulse governing ventricular contraction from the AV node to the ventricular apex. Disorders of the VCS are common, carry significant morbidity, and are poorly understood from a molecular perspective.The transcriptional networks required to maintain function of the adult CCS are undefined. Our current understanding of the molecular mediators of CCS function stems largely from heritable monogenic disorders and mouse models that have identified a limited number of genes essential for maintaining cardiac rhythm, most of which encode ion channels and their interacting partners (reviewed in ref. 1). Similar approaches have also begun to uncover the transcriptional networks required for CCS development (reviewed in ref.2). Recent genome-wide association studies (GWAS) have identified loci implicated in ECG interval variation (3-8), providing candidate genes with potentially important roles in CCS function in the general population. Specifically, numerous loci near genes e...

A common genetic variant within SCN10A modulates cardiac SCN5A expression

“…Expression analysis also supports a more substantial role for the gene product of Scn5a than that of Scn10a. During embryonic development, low levels of Scn10a expression can be qualitatively observed using in situ hybridization (21,22) and qRT-PCR, demonstrating expression levels 4-5 times lower than those of Scn5a. However, as development progressed to adult stages, Scn10a expression remained low, whereas Scn5a transcript levels were induced to levels 100-fold greater than those of Scn10a.…”

“…expression of a reporter construct in the murine CCS (21,22). The region termed enhancer A (EnhA) spans 2 adjacent introns and 1 exon in Scn10a, and enhancer B (EnhB) is positioned 15 kbp downstream of Scn5a ( Figure 1B).…”

“…We recently demonstrated the importance of the T-box transcription factors Tbx5 and Tbx3 in the regulation of Scn5a in mice (21,22). The rapid conducting portions of the CCS require high levels of Na v 1.5 for function (23).…”

“…The rapid conducting portions of the CCS require high levels of Na v 1.5 for function (23). Strong Scn5a expression is driven in the fast conduction system by TBX5, a transcriptional activator (22). Conversely, repression of Scn5a is a key feature ensuring the slow propagation of the cardiac electrical impulse across the sinus node and atrioventricular node (17,23).…”

The formation and function of the cardiac conduction system

“…Hcn4). Tbx5 is expressed more broadly in the ventricle, but in the AVB and BB it stimulates the gene programme for fast conduction, including Cx40 and Scn5a (Moskowitz et al, 2004;Arnolds et al, 2012). Here, it successfully competes with Tbx3 for occupation of binding sites of 'conduction genes' such as Cx40 and Scn5a.…”