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Cardiovascular Research 2004 61(1):56-65; doi:10.1016/j.cardiores.2003.09.030
© 2004 by European Society of Cardiology
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Copyright © 2004, European Society of Cardiology

Cloning and initial characterization of the human cardiac sodium channel (SCN5A) promoter *

Ping Yanga, Sabina Kupershmidta,b and Dan M. Roden*,a,c

aDepartment of Pharmacology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA
bDepartment of Anesthesiology, Vanderbilt University School of Medicine, USA
cDepartment of Medicine, Vanderbilt University School of Medicine, USA

* Corresponding author. Division of Clinical Pharmacology, Department of Pharmacology, Vanderbilt University School of Medicine, 532 Medical Research Building I, Nashville, TN 37232, USA. Tel.: +1-615-322-0067; fax: +1-615-343-4522. dan.roden{at}vanderbilt.edu

Objective: Despite the primacy of the sodium current in cardiac electrophysiology and evidence that decreased sodium current is arrhythmogenic in humans, little is known about transcriptional regulation of the underlying gene, SCN5A. Methods: We have cloned a 2.7 kb segment of 5'-flanking region of SCN5A and identified multiple transcription initiation sites by primer extension and RNase protection. Transient transfection assays in neonatal mouse myocytes and in Chinese Hamster Ovary (CHO) cells were employed to identify promoter activities. PCR-single stranded conformational polymorphism (SSCP) analysis was used to screen DNA variants in the promoter region. Results: The fragment includes >2 kb of upstream sequence, the 173-bp non-coding exon 1, and a portion of the 16-kb intron 1; the region is highly GC-rich and TATA-less. Transient transfection assays in neonatal mouse myocytes and in CHO cells identified (1) a core promoter in the –261/+140 segment, (2) regions conferring ~3-fold decreases from core promoter activity in the 5' upstream region (–261/–454 and –1020/–2109), and ~3-fold increases in intron 1 (+255/+410 and+539/+613), and (3) a very strong negative regulatory region between +613 and +754 in intron 1. A core promoter polymorphism, present in 6/142 (4%) of normal alleles screened, increased reporter activity ~50% in myocytes but not in CHO cells. Conclusion: The SCN5A promoter includes multiple positive and negative cis-acting elements extending into intron 1. A common polymorphism in this region modulates channel expression in vitro.

KEYWORDS Ion channels; Arrhythmia (mechanism); Gene polymorphisms

Time for primary review 24 days

* The nucleotide sequence for the human hSCN5A promoter has been deposited in the GenBank database under GenBank accession number AY313163. The nucleotide sequence for the mouse mscn5a promoter has been deposited in the GenBank database under GenBank accession number AY313164.


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