Pitx2 modulates a Tbx5-dependent gene regulatory network to maintain atrial rhythm.

Rangarajan D Nadadur,Yun Qiao,Margaret Gadek,James F Martin,Xinan Yang,Bastiaan Boukens,Stefan R Mazurek,Malou Van Den Boogaard,Min Zhang,Elizabeth M Mcnally,Jon Seidman,Vincent Christoffels,Igor R Efimov,Tarsha Ward,Christopher R Weber,Christine E Seidman,Michael T Broman,Jenna Bekeny,Ivan P Moskowitz

doi:10.1126/scitranslmed.aaf4891

Abstract

Cardiac rhythm is extremely robust, generating 2 billion contraction cycles during the average human life span. Transcriptional control of cardiac rhythm is poorly understood. We found that removal of the transcription factor gene Tbx5 from the adult mouse caused primary spontaneous and sustained atrial fibrillation (AF). Atrial cardiomyocytes from the Tbx5-mutant mice exhibited action potential abnormalities, including spontaneous depolarizations, which were rescued by chelating free calcium. We identified a multitiered transcriptional network that linked seven previously defined AF risk loci: TBX5 directly activated PITX2, and TBX5 and PITX2 antagonistically regulated membrane effector genes Scn5a, Gja1, Ryr2, Dsp, and Atp2a2 In addition, reduced Tbx5 dose by adult-specific haploinsufficiency caused decreased target gene expression, myocardial automaticity, and AF inducibility, which were all rescued by Pitx2 haploinsufficiency in mice. These results defined a transcriptional architecture for atrial rhythm control organized as an incoherent feed-forward loop, driven by TBX5 and modulated by PITX2. TBX5/PITX2 interplay provides tight control of atrial rhythm effector gene expression, and perturbation of the co-regulated network caused AF susceptibility. This work provides a model for the molecular mechanisms underpinning the genetic implication of multiple AF genome-wide association studies loci and will contribute to future efforts to stratify patients for AF risk by genotype.

Highlights

The transcriptional architecture that confers robustness to cardiac rhythm must tightly control cardiac channel gene expression, because increased or decreased channel expression can cause cardiac arrhythmias [1]
Tbx5 was deleted from the adult mouse by combining a Tbx5 floxed allele (Tbx5fl) [5] with a tamoxifen (TM)–inducible Cre recombinase allele at the Rosa26 (R26) locus [14]
ECGs of ambulatory TM-treated Tbx5fl/fl; R26CreERt2 mice showed a disorganized pattern of atrial activity compared to TM-treated R26CreERt2 controls (Fig. 1A)

Summary

Introduction

The transcriptional architecture that confers robustness to cardiac rhythm must tightly control cardiac channel gene expression, because increased or decreased channel expression can cause cardiac arrhythmias [1]. Human AF GWAS have implicated multiple transcription factors, including TBX5 and PITX2 in AF, raising the possibility that perturbations of a gene regulatory network for atrial rhythm control may underlie some AF susceptibility [3]. Dominant mutations in the T-box transcription factor TBX5 cause Holt-Oram syndrome, characterized by disrupted heart and limb development [4,5,6] and increased AF risk [6]. PITX2, a paired-like homeodomain transcription factor, plays a critical role in heart development and adult rhythm control. Adult-specific Pitx deletion in mice causes AF susceptibility and increased expression of ion channels linked to AF [11]

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