Journal article
The structure of the rat amiloride-sensitive epithelial sodium channel gamma subunit gene and functional analysis of its promoter
Gene, Vol.228(1), pp.111-122
1999
DOI: 10.1016/S0378-1119(99)00016-5
PMID: 10072764
Abstract
Prolonged dietary Na
+ depletion and chronic administration of adrenal steroids increase steady-state mRNA levels of the γ subunit of the epithelial sodium channel (γENaC) in rat colon. This increase correlates with a marked increase in transepithelial Na
+ transport and is thought to occur via transcriptional regulation. To begin to evaluate these mechanisms in detail, we determined the organization of the rat γENaC gene. A rat genomic library was screened and overlapping lambda clones that together span the gene (∼36
kb) and contain at least 1
kb of 5′ flanking genomic DNA were isolated. As in the human gene, the rat γENaC gene contains 13 exons and a CpG island at the 5′ end of the gene. A single transcription start site was identified in rat kidney by nuclease protection assay defining a 5′ untranslated region of 126
nt. The translation initiation codon was identified within the second exon and the entire 3′ UTR (∼1
kb) was within the last exon. 800
bp of 5′ flanking sequence, as well as the 3.4
kb first intron, were sequenced and analyzed for transcriptional regulatory motifs. Analogous to the human γENaC gene [Thomas, C.P., Doggett, N.A., Fisher, R., Stokes J.B., 1996. Genomic organization and the 5′ flanking region of the gamma subunit of the human amiloride-sensitive epithelial sodium channel. J. Biol. Chem. 271, 26
062–26
066], two GC boxes were seen at −30 and −61 to the transcription start site. In addition, putative AP-1, AP-2, CRE, Sp1 and GATA-1 and GRE motifs were identified elsewhere in the 5′ flanking region or the first intron. Two mammalian-wide interspersed repeats and a rodent-specific B1 repeat were also identified within the first intron. Fragments containing the putative GRE motifs coupled to luciferase did not confer a glucocorticoid-stimulated response in two cell lines that contained a functional glucocorticoid receptor. However, a 76
nt rat γENaC 5′ flanking fragment (−76 to +68) directed expression of luciferase in the epithelial cell lines H441 and FRTL5, suggesting that this minimal region that contained both GC boxes was sufficient for promoter activity.
Details
- Title: Subtitle
- The structure of the rat amiloride-sensitive epithelial sodium channel gamma subunit gene and functional analysis of its promoter
- Creators
- Christie P Thomas - Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, IA, USAScott D Auerbach - Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, IA, USAChong ZhangJohn B Stokes - Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, IA, USA
- Resource Type
- Journal article
- Publication Details
- Gene, Vol.228(1), pp.111-122
- Publisher
- Elsevier B.V
- DOI
- 10.1016/S0378-1119(99)00016-5
- PMID
- 10072764
- ISSN
- 0378-1119
- eISSN
- 1879-0038
- Language
- English
- Date published
- 1999
- Academic Unit
- Stead Family Department of Pediatrics; Obstetrics and Gynecology; Nephrology; Internal Medicine
- Record Identifier
- 9983985903402771
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