Acidification and glucocorticoids independently regulate branched-chain α-ketoacid dehydrogenase subunit genes

Abstract

Acidification or glucocorticoids increase the maximal activity and subunit mRNA levels of branched chain α-ketoacid dehydrogenase (BCKAD) in various cell types. We examined whether these stimuli increase transcription of BCKAD subunit genes by transfecting BCKAD subunit promoter-luciferase plasmids containing the mouse E2 or human E1α-subunit promoter into LLC-PK1 cells, which do not express glucocorticoid receptors, or LLC-PK1-GR101 cells, which we have engineered to constitutively express the glucocorticoid receptor gene. Dexamethasone or acidification increased luciferase activity in LLC-PK1-GR101 cells transfected with the E2 or E1α-minigenes; acidification augmented luciferase activity in LLC-PK1 cells transfected with these minigenes but dexamethasone did not. A pH-responsive element in the E2 subunit promoter was mapped to a region >4.0 kb upstream of the transcription start site. Dexamethasone concurrently stimulated E2 subunit promoter activity and reduced the binding of nuclear factor-κB (NF-κB) to a site in the E2 promoter. Thus acidification and glucocorticoids independently enhance BCKAD subunit gene expression, and the glucocorticoid response in the E2 subunit involves interference with NF-κB, which may act as a transrepressor.