Promoter DNA Methylation Regulates Murine SUR1 (Abcc8) and SUR2 (Abcc9) Expression in HL-1 Cardiomyocytes

Abstract

Two mammalian genes encode the SURx (SUR1, Abcc8 and SUR2, Abcc9) subunits that combine with Kir6.2 (Kcnj11) subunits to form the ATP-sensitive potassium (KATP) channel in cardiac myocytes. Different isoform combinations endow the channel with distinct physiological and pharmacological properties, and we have recently reported that the molecular composition of sarcolemmal KATP channels is chamber specific in the mouse heart. KATP channel composition is determined by what subunits are expressed in a cell or tissue. In the present study, we explore the role of CpG methylation in regulating SUR1 and SUR2 expression. In HL-1 cardiomyocytes, as in atrial myocytes, SUR1 expression is markedly greater than SUR2. Consistent with CpG methylation-dependent silencing of SUR2 expression, bisulfite sequencing of genomic DNA isolated from HL-1 cells demonstrates that 57.6% of the CpGs in the promoter region of the SUR2 gene are methylated, compared with 0.14% of the the CpG residues in the SUR1 sequence. Moreover, treatment with 10 µM 5-aza-2′-deoxycytidine (Aza-dC) significantly increased both the unmethylated fraction of the SUR2 CpG island and mRNA expression. However, we cannot rule out additional mechanisms of Aza-dC action, as Aza-dC also causes a decrease in SUR1 expression and lower doses of Aza-dC do not alter the unmethylated DNA fraction but do elicit a small increase in SUR2 expression. The conclusion that DNA methylation alone is not the only regulator of SUR subunit expression is also consistent with observations in native myocytes, where the CpG islands of both SUR genes are essentially unmethylated in both atrial and ventricular myocytes. Collectively, these data demonstrate the potential for CpG methylation to regulate SURx subunit expression and raises the possibility that regulated or aberrant CpG methylation might play a role in controlling channel structure and function under different physiological conditions or different species.

KATPChannels in the Cardiovascular System

“…Transcriptional regulation of the K ATP channel subunit genes (KCNJ8, KCNJ11, ABCC8, and ABCC9) has not been studied systematically. A recent study showed that the ABCC8 and ABCC9 promoters can be methylated in HL-1 cells and that methylation decreases mRNA expression (220). Although these genes were not methylated in the mouse heart, the possibility is raised that DNA methylation may regulate transcriptional activity of K ATP channel genes.…”

Mechanisms contributing to myocardial potassium channel diversity, regulation and remodeling

“…The protein expression levels of KChIP2 ( Kcnip2 ), but not Kv4.2 ( Kcnd2 ) or Kv4.3 ( Kcnd3 ), were reported to be reduced significantly in PTIP −/− mouse hearts, leading to decreased I to,f densities and action potential prolongation, as well as impaired calcium handling and contractility [55]. Promoter DNA methylation modulates the expression of the I KATP channel subunits, SUR1 and SUR2, in mouse cardiomyocytes [56]. The extent of promoter CpG methylation inversely correlates with the expression of SUR1 and SUR2, and treatment with the DNA methylation inhibitor 5′-Aza-2′ deoxycytidine significantly reduced both methylation at the SUR2 CpG island and SUR2A mRNA expression [56].…”

ABCC9/SUR2 in the brain: Implications for hippocampal sclerosis of aging and a potential therapeutic target

“…Interestingly, the human SUR2 3’UTR is variable in length (al) which may be a pathway for regulating transcript stability (Nelson and Keller, 2007). Further, mouse Sur1 and Sur2 expression can be regulated through promoter DNA methylation (Fatima, et al, 2012). …”