POPDC2 a novel susceptibility gene for conduction disorders

Abstract

Despite recent progress in the understanding of cardiac ion channel function and its role in inherited forms of ventricular arrhythmias, the molecular basis of cardiac conduction disorders often remains unresolved. We aimed to elucidate the genetic background of familial atrioventricular block (AVB) using a whole exome sequencing (WES) approach. In monozygotic twins with a third-degree AVB and in another, unrelated family with first-degree AVB, we identified a heterozygous nonsense mutation in the POPDC2 gene causing a premature stop at position 188 (POPDC2^W188⁎), deleting parts of its cAMP binding-domain. Popeye-domain containing (POPDC) proteins are predominantly expressed in the skeletal muscle and the heart, with particularly high expression of POPDC2 in the sinoatrial node of the mouse. We now show by quantitative PCR experiments that in the human heart the POPDC-modulated two-pore domain potassium (K_2P) channel TREK-1 is preferentially expressed in the atrioventricular node. Co-expression studies in Xenopus oocytes revealed that POPDC2^W188⁎ causes a loss-of-function with impaired TREK-1 modulation. Consistent with the high expression level of POPDC2 in the murine sinoatrial node, POPDC2^W188⁎ knock-in mice displayed stress-induced sinus bradycardia and pauses, a phenotype that was previously also reported for POPDC2 and TREK-1 knock-out mice. We propose that the POPDC2^W188⁎ loss-of-function mutation contributes to AVB pathogenesis by an aberrant modulation of TREK-1, highlighting that POPDC2 represents a novel arrhythmia gene for cardiac conduction disorders.

Introduction

Disorders affecting the cardiac conduction system can induce serious and potentially life-threatening bradyarrhythmias. These dysfunctions can become evident by impaired cardiac conduction through the atrioventricular (AV) node, the His-bundle or the Purkinje system with right or left bundle branch block (RBBB or LBBB) and/or a broadening of the QRS complex. Hereditary forms of arrhythmias are predominantly caused by mutations in genes encoding ion channels or their modulating subunits. Mutations in the SCN5A gene encoding the cardiac sodium channel or its β-subunits are associated with progressive cardiac conduction disorder (PCCD) [[1], [2], [3]] and with congenital sick sinus syndrome (SSS) [1,4]. In addition, mutations in the TRPM4 channel gene cause autosomal-dominant hereditary forms of PCCD including AV conduction abnormalities [5]. Moreover, mutations in the LMNA gene may first present as AV conduction disorder and later as progressive heart failure and dilated cardiomyopathy [6]. Despite recent progress in the understanding of cardiac ion channel function and their role in inherited forms of ventricular arrhythmias, the genetic background of cardiac conduction disorders often remains unresolved.

The Popeye domain containing (POPDC) gene family encodes proteins with three transmembrane domains which are predominantly expressed in the heart and skeletal muscle [7,8]. The C-terminal parts of the proteins harboring the highly conserved Popeye-domain which contains a cAMP binding site are localized to the cytoplasm [9]. POPDC proteins are known to interact with the two-pore domain potassium (K_2P) channel TREK-1 thereby enhancing its current amplitude [9]. We have previously reported that cAMP binds with high affinity to the Popeye domain, abolishing the current modulation of TREK-1 channels [9].

TREK-1 plays a major role in murine sinoatrial pacemaking as cardiac-specific ablation of TREK-1 in mice causes a bradycardia with episodes of sinus pauses following stress [10]. In the mouse heart, POPDC2 is expressed in atrial and ventricular myocytes [8], however a dominant expression in the sinoatrial and AV node and other parts of the cardiac conduction system was described [9]. Interestingly, in mice, POPDC1 and POPDC2 null mutants presented a comparable phenotype as the TREK-1 knock-out mice, with a strong stress-induced sinoatrial node dysfunction [9,10]. On the other hand, in POPDC1/POPDC2 double knock-out mice also an AVB was prevalent [11], compatible with observations from zebrafish in which morpholino oligonucleotide-mediated knock-down of POPDC2 led to AVB [12]. In humans we recently identified a POPDC1^S201F mutation which causes AV block (AVB) and autosomal recessive limb-girdle muscular dystrophy (LGMDR25) [13]. Another three recessive mutations in POPDC1 are associated with AVB and LGMD of variable severity and age at onset [14], while mutations in POPDC3 are causing LGMD (LGMDR26) with no cardiac involvement [15].

To elucidate the genetic background of familial AVB we used a whole exome sequencing (WES) approach in a family with AVB and identified the heterozygous POPDC2 nonsense variant POPDC2^W188⁎. The same POPDC2^W188⁎ mutation was subsequently identified in another family with dominant inheritance. Mechanistically, our data suggest that the POPDC2^W188⁎ mutation causes impaired AV conduction that is mediated by a dysregulation of TREK-1 potassium channels.