Genomic organization of the 5' end of human beta-ENaC and preliminary characterization of its promoter

The mRNA for the beta-subunit of the epithelial Na(+) channel (beta-ENaC) is regulated developmentally and, in some tissues, in response to corticosteroids. To understand the mechanisms of transcriptional regulation of the human beta-ENaC gene, we characterized the 5' end of the gene and its 5&...

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Bibliographic Details
Published in:American journal of physiology. Renal physiology 2002-05, Vol.282 (5), p.F898-F909
Main Authors: Thomas, Christie P, Loftus, Randy W, Liu, Kang Z, Itani, Omar A
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Base Sequence
Cloning, Molecular
Cosmids
DNA, Complementary - analysis
Epithelial Sodium Channels
Exons
Gene Amplification
Gene Expression Regulation
Gene Library
Humans
Immunoblotting
Kidney - chemistry
Lung - chemistry
Molecular Sequence Data
Promoter Regions, Genetic
Protein Biosynthesis
RNA, Messenger - analysis
Sodium Channels - chemistry
Sodium Channels - genetics
Sp1 Transcription Factor - metabolism
Transcription, Genetic
Transfection
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Summary:The mRNA for the beta-subunit of the epithelial Na(+) channel (beta-ENaC) is regulated developmentally and, in some tissues, in response to corticosteroids. To understand the mechanisms of transcriptional regulation of the human beta-ENaC gene, we characterized the 5' end of the gene and its 5'-flanking regions. Adaptor-ligated human kidney and lung cDNA were amplified by 5' rapid amplification of cDNA ends, and transcription start sites of two 5' variant transcripts were determined by nuclease protection or primer extension assays. Cosmid clones that contain the 5' end of the gene were isolated, and analysis of these clones indicated that alternate first exons approximately 1.5 kb apart and approximately 45 kb upstream of a common second exon formed the basis of these transcripts. Genomic fragments that included the proximal 5'-flanking region of either transcript were able to direct expression of a reporter gene in lung epithelia and to bind Sp1 in nuclear extracts, confirming the presence of separate promoters that regulate beta-ENaC expression.
ISSN:1931-857X
1522-1466
DOI:10.1152/ajprenal.00268.2001