Identification of Key Small Non‐Coding MicroRNAs Controlling Pacemaker Mechanisms in the Human Sinus Node

Abstract

Background The sinus node (SN) is the primary pacemaker of the heart. SN myocytes possess distinctive action potential morphology with spontaneous diastolic depolarization because of a unique expression of ion channels and Ca 2+ ‐handling proteins. MicroRNAs (miRs) inhibit gene expression. The role of miRs in controlling the expression of genes responsible for human SN pacemaking and conduction has not been explored. The aim of this study was to determine miR expression profile of the human SN as compared with that of non‐pacemaker atrial muscle. Methods and Results SN and atrial muscle biopsies were obtained from donor or post‐mortem hearts (n=10), histology/immunolabeling were used to characterize the tissues, TaqMan Human MicroRNA Arrays were used to measure 754 miRs, Ingenuity Pathway Analysis was used to identify miRs controlling SN pacemaker gene expression. Eighteen miRs were significantly more and 48 significantly less abundant in the SN than atrial muscle. The most interesting miR was miR‐486‐3p predicted to inhibit expression of pacemaking channels: HCN1 (hyperpolarization‐activated cyclic nucleotide‐gated 1), HCN4, voltage‐gated calcium channel (Ca v )1.3, and Ca v 3.1. A luciferase reporter gene assay confirmed that miR‐486‐3p can control HCN4 expression via its 3′ untranslated region. In ex vivo SN preparations, transfection with miR‐486‐3p reduced the beating rate by ≈35±5% ( P <0.05) and HCN4 expression ( P <0.05). Conclusions The human SN possesses a unique pattern of expression of miRs predicted to target functionally important genes. miR‐486‐3p has an important role in SN pacemaker activity by targeting HCN4, making it a potential target for therapeutic treatment of SN disease such as sinus tachycardia.

A detailed characterization of the hyperpolarization-activated “funny” current (If) in human-induced pluripotent stem cell (iPSC)–derived cardiomyocytes with pacemaker activity

“…Figure 2d shows a representative immunofluorescence image of hiPSC-CMs co-stained with anti-HCN4 and -caveolin-3 antibodies. As previously demonstrated, the co-expression of these two proteins is characteristic of pacemaker/SAN cardiomyocytes of different species [ 4 , 33 , 36 ].…”

Post-Transcriptional Regulation of Molecular Determinants during Cardiogenesis

“…Wang et al [ 185 ] demonstrated that Pitx2 positively regulates miR-17-92 and miR-106b-25 repressing the sinoatrial node genetic program ( Figure 1 B–D). On the other hand, several studies have evidenced that Pitx2 modulates the expression of several microRNAs that contribute to Atrial Fibrillation (AF), such as miR-106-25, miR-17-92, miR-29a, miR-200, miR-203, miR-21, miR-208ab, miR-1, miR-26b and miR-106ab [ 185 , 188 , 189 , 190 , 191 , 192 ]. The lack of Pitx2 expression in the left atrial appendage impairs the expression of a microRNA battery (miR-21, miR-106a, miR-203 and miR-208ab down-regulated, and miR-1, miR-26b, miR-29a, miR-106b and miR-200 up-regulated) through a Wnt8a and Wnt11 modulation, leading to a calcium, sodium and potassium channel remodeling during AF [ 185 , 188 , 189 ].…”

Inherited and Acquired Rhythm Disturbances in Sick Sinus Syndrome, Brugada Syndrome, and Atrial Fibrillation: Lessons from Preclinical Modeling

“…Finally, extensive linkage and genome-wide associated studies [18,[45][46][47][48][49][50][51][52], posttranscriptional analyses, and next-generation sequencing [49,[53][54][55][56][57][58][59][60][61][62] on animal and human tissues and cells, included pluripotent stem cells [63,64], shed light on the ontology of the genes regulated during CCS development and physiology, as well as on their fine-tune silencing provided by specific microRNAs (e.g., miR-1 and miR-128a). Each component of the CCS is, hence, the developmental embryology result of unique hierarchical transcriptional networks, with a prominent orchestrating or ruling role by T-box transcription factors, as previously summarized by Munshi in 2012 [65].…”