Transcriptional control of pacemaker channel genes HCN2 and HCN4 by Sp1 and implications in re-expression of these genes in hypertrophied myocytes.

Cardiac hypertrophy is characterized by electrical remolding with increased risk of arrhythmogenesis. Enhanced abnormal automaticity of ventricular cells may contribute to hypertrophic arrhythmias. The pacemaker current I(f), carried by the hyperpolarization-activated channels encoded mainly by the HCN2 and HCN4 genes in the heart, plays an important role in rhythmogenesis. Their expressions reportedly increase in hypertrophic and failing hearts, contributing to arrhythmogenesis under these conditions. However, how their expressions are controlled remained unclear. We performed a study to characterize the regulatory elements and transcriptional control of HCN2 and HCN4 genes. We identified the transcription start sites by 5'RACE and core promoter regions of these genes using luciferase reporter assay, and revealed the ubiquitous Sp1 protein as a common transactivator of HCN2 and HCN4 genes. We further unraveled robust increases in HCN2/HCN4 transcripts and protein levels, using real-time RT-PCR and Western blot analyses, in a rat model of left ventricular hypertrophy and in angiotensin II-induced neonatal ventricular hypertrophy. The upregulation of HCN2 and HCN4 transcription was accompanied by pronounced elevations of Sp1 and silencing of Sp1 by siRNA prevented overexpression of HCN2/HCN4 in hypertrophic cardiomyocytes. Our data indicate that Sp1 drives HCN2/HCN4 transcription and determines the functional level of HCN2/HCN4 mRNAs, and upregulation of Sp1 underlie the abnormal re-expression of HCN2/HCN4 genes in hypertrophied myocytes. This study also provides the first evidence for the role of Sp1 in the reactivation of 'fetal' cardiac genes, HCN2 and HCN4, in ventricular myocytes, and thereby in the pathological electrical remodeling in hypertrophied myocytes.