title: Glucocorticoids can activate the alpha-ENaC gene promoter independently of SGK1.
abstract: The role of SGK1 (serum- and glucocorticoid-induced protein kinase 1) in the glucocorticoid induction of alpha-ENaC (epithelial Na+ channel alpha subunit) gene transcription was explored by monitoring the transcriptional activity of a luciferase-linked, alpha-ENaC reporter gene construct (pGL3-KR1) expressed in H441 airway epithelial cells. Dexamethasone evoked a concentration-dependent (EC50 approximately 4 microM) increase in transcriptional activity dependent upon a glucocorticoid response element in the alpha-ENaC sequence. Although dexamethasone also activated endogenous SGK1, artificially increasing cellular SGK1 activity by expressing a constitutively active SGK1 mutant (SGK1-S422D) in hormone-deprived cells did not activate pGL3-KR1. Moreover, expression of catalytically inactive SGK1 (SGK1-K127A) suppressed the activation of endogenous SGK1 without affecting the transcriptional response to dexamethasone. Increasing cellular PI3K (phosphoinositide 3-kinase) activity by expressing a membrane-anchored form of the catalytic PI3K-P110alpha subunit [CD2 (cluster of differentiation 2)-P110alpha] also activated endogenous SGK1 without affecting pGL3-KR1activity. A catalytically inactive form of CD2-P110alpha (R1130P), on the other hand, prevented the dexamethasone-induced activation of SGK1, but did not inhibit the activation of pGL3-KR1. However, expression of SGK1-S422D or CD2-P110alpha enhanced the transcriptional responses to maximally effective concentrations of dexamethasone and this effect occurred with no change in EC50. Dexamethasone-induced (0.3-300 nM) activation of pGL3-KR1 was unaffected by inhibitors of PI3K (PI-103 and wortmanin) and by rapamycin, a selective inhibitor of the TORC1 (target of rapamycin complex 1) signalling complex. Dexamethasone-induced activation of the alpha-ENaC gene promoter can thus occur independently of SGK1/PI3K, although this pathway does provide a mechanism that allows this transcriptional response to dexamethasone to be enhanced.
authors.affiliation: Centre for Cardiovascular and Pulmonary Research, Division of Medical Sciences, Ninewells Hospital and Medical School, University of Dundee, Dundee, UK.
authors.forename: Niall
authors.initials: N
authors.lastname: McTavish
authors.forename: Jennet
authors.initials: J
authors.lastname: Getty
authors.forename: Ann
authors.initials: A
authors.lastname: Burchell
authors.forename: Stuart M
authors.initials: SM
authors.lastname: Wilson
mesh_terms: D018797:Cell Cycle Proteins / Q000378:metabolism; D002478:Cells, Cultured; D003907:Dexamethasone / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D053503:Epithelial Sodium Channels / Q000235:genetics*; D005938:Glucocorticoids / Q000494:pharmacology*; D006801:Humans; D017874:Immediate-Early Proteins / Q000378:metabolism / Q000502:physiology*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation / Q000187:drug effects; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism / Q000502:physiology*; D013312:Stress, Physiological / Q000502:physiology; D015533:Transcriptional Activation / Q000187:drug effects; D014162:Transfection; D000099184:Serum-Glucocorticoid Regulated Kinases; D000099259:N-myc Downstream-Regulated Gene 1 Protein
chemical_list: D018797:Cell Cycle Proteins; D003907:Dexamethasone; D004791:Enzyme Inhibitors; D053503:Epithelial Sodium Channels; D005938:Glucocorticoids; D017874:Immediate-Early Proteins; D047908:Intracellular Signaling Peptides and Proteins; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases; D000099184:Serum-Glucocorticoid Regulated Kinases; D000099259:N-myc Downstream-Regulated Gene 1 Protein