Hormonal regulation and genomic organization of the human amiloride-sensitive epithelial sodium channel alpha subunit gene.
To investigate the regulation of the amiloride-sensitive epithelial sodium channel (ENaC) expression, we have characterized the genomic structure and performed promoter analyses of the alpha subunit of the human (h) ENaC gene. Genomic clones containing the alphahENaC gene were isolated and subjected to restriction-mapping analysis. The alphahENaC gene was shown to be composed of 13 exons and 12 introns. Primer extension analysis confirmed that transcription initiation occurred at the beginning of the reported alphahENaC cDNA, but also indicated potential heterogenous initiation sites. Examination of a 3.1 kb 5' flanking sequence revealed a notable absence of CCAAT or TATA-like elements but suggested three GC boxes and several putative transcription factor binding sites, including a glucocorticoid response element (GRE) consensus. A 250 bp minimal promoter was capable of directing expression of a secreted alkaline phosphatase reporter. This promoter activity was enhanced 2.5- and 4-fold by upstream flanking sequences. Dexamethasone treatment induced levels of expression from the longer, GRE-containing promoter fragments from 8- to 20-fold, but not from the minimal promoter. Precise deletion of the 15-bp, dyad GRE sequence completely abolishes the response of reporter expression to dexamethasone induction. These experiments indicate that glucocorticoid augmentation of lung epithelial Na+ transport occurs, at least in part, by direct stimulation of transcription of the ENaC genes.
Chow
Y H
YH
Programme in Lung Biology Research, Hospital for Sick Children, Toronto, Ontario, Canada.
Wang
Y
Y
Plumb
J
J
O'Brodovich
H
H
Hu
J
J
D001483:Base Sequence; D018076:DNA, Complementary / Q000032:analysis / Q000235:genetics; D003907:Dexamethasone / Q000494:pharmacology; D053503:Epithelial Sodium Channels; D005786:Gene Expression Regulation / Q000187:drug effects; D015894:Genome, Human; D005938:Glucocorticoids / Q000494:pharmacology; D006801:Humans; D017136:Ion Transport; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D015222:Sodium Channels / Q000235:genetics*
D018076:DNA, Complementary; D053503:Epithelial Sodium Channels; D005938:Glucocorticoids; D015222:Sodium Channels; D003907:Dexamethasone