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'-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.
Thomas
Christie P
CP
Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, Iowa 52242-1081, USA. christie-thomas@uiowa.edu
Loftus
Randy W
RW
Liu
Kang Z
KZ
Itani
Omar A
OA
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D003360:Cosmids; D018076:DNA, Complementary / Q000032:analysis; D053503:Epithelial Sodium Channels; D005091:Exons; D005784:Gene Amplification; D005786:Gene Expression Regulation; D015723:Gene Library; D006801:Humans; D015151:Immunoblotting; D007668:Kidney / Q000737:chemistry; D008168:Lung / Q000737:chemistry; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000032:analysis; D015222:Sodium Channels / Q000737:chemistry / Q000235:genetics*; D016329:Sp1 Transcription Factor / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection
D018076:DNA, Complementary; D053503:Epithelial Sodium Channels; D012333:RNA, Messenger; D015222:Sodium Channels; D016329:Sp1 Transcription Factor