RNA Sequencing of Mouse Sinoatrial Node Reveals an Upstream Regulatory Role for Islet-1 in Cardiac Pacemaker Cells

Abstract

Rationale Treatment of sinus node disease with regenerative or cell-based therapies will require a detailed understanding of gene regulatory networks in cardiac pacemaker cells (PCs). Objective To characterize the transcriptome of PCs using RNA sequencing, and to identify transcriptional networks responsible for PC gene expression. Methods and Results We used laser capture micro-dissection (LCM) on a sinus node reporter mouse line to isolate RNA from PCs for RNA sequencing (RNA-Seq). Differential expression and network analysis identified novel SAN-enriched genes, and predicted that the transcription factor Islet-1 (Isl1) is active in developing pacemaker cells. RNA-Seq on SAN tissue lacking Isl1 established that Isl1 is an important transcriptional regulator within the developing SAN. Conclusions (1) The PC transcriptome diverges sharply from other cardiomyocytes; (2) Isl1 is a positive transcriptional regulator of the PC gene expression program.

The formation and function of the cardiac conduction system

“…Recently, the LIM-homeobox transcription factor Isl1 has also gained attention as an important regulator of SAN development (Hoffmann et al, 2013;Liang et al, 2015;Vedantham et al, 2015). Isl1 is transiently expressed in, and is required for, cardiac mesodermal progenitors, and is downregulated as soon as they differentiate to cardiomyocytes.…”

“…Isl1 is transiently expressed in, and is required for, cardiac mesodermal progenitors, and is downregulated as soon as they differentiate to cardiomyocytes. However, its expression is selectively maintained in myocytes of the SAN, both during embryogenesis and in the adult (Sun et al, 2007;Mommersteeg et al, 2010;Sizarov et al, 2011;Weinberger et al, 2012;Vedantham et al, 2015;Liang et al, 2015). In zebrafish, Isl1 marks pacemaker cells in the junction of the sinus venosus and atrium, and it is required for normal pacemaker function and development (Tessadori et al, 2012).…”

“…In mouse, Isl1 is required for the proliferation and function of SAN cells, and the SAN-specific deletion of Isl1 results in embryonic lethality . Furthermore, Isl1-deficiency in mice leads to a downregulation of key regulators of SAN development, such as Tbx3, Shox2 and Bmp4, and ion channels for SAN function, including Hcn4, Hcn1 and Cacna1g Vedantham et al, 2015), whereas Isl1 overexpression in ESC-derived cardiomyocytes leads to upregulation of the SAN gene programme and downregulation of the chamber myocardium gene programme (Dorn et al, 2015). Isl1 is a target of Shox2 in the SAN and can rescue the bradycardia phenotype caused by Shox2 deficiency (Hoffmann et al, 2013).…”

Transcription factor ISL1 is essential for pacemaker development and function

“…Due to early lethality and loss of cardiac cells derived from ISL1 progenitors in Isl1-null mice, a specific role for ISL1 in the SAN remains unknown. Recent transcriptome studies demonstrated changes in gene expression in response to Isl1 ablation in the SAN (60). However, no studies have been performed to address phenotypic or physiological consequences of Isl1 ablation.…”

Human fetal cardiac progenitors: The role of stem cells and progenitors in the fetal and adult heart

“…However, Isl1 protein has been detected already at the cardiac crescent stage, suggesting the possibility that Isl1 is expressed in FHF at a very early stage [56]. In the OFT of the human fetal hearts at gestation week 9, Lui and colleagues identified an Isl1-positive population of endothelial progenitors expressing VEGF-A receptors [57], whereas Vedantham et al demonstrated that Isl1 is a regulator of sinoatrial node development [58]. Thus, in contrast to HCN4, Isl1 expression may identify a transient progenitor state important for heart lineage diversification that rapidly converges into the vascular endothelial cell, myocardial, and conduction system cell lineages, followed by downregulation of Isl1 upon cardiomyocyte differentiation [53,55].…”