Down-regulation of rat mitochondrial branched-chain 2-oxoacid dehydrogenase kinase gene expression by glucocorticoids.
The mammalian mitochondrial branched-chain 2-oxoacid dehydrogenase (BCOD) complex is regulated by a reversible phosphorylation (inactivation)/dephosphorylation (activation) cycle. In the present study, the effects of glucocorticoids on the level of BCOD kinase mRNA were investigated in rat hepatoma cell lines (H4IIE and FTO-2B), as well as in the rat. In H4IIE cells, dexamethasone was found to significantly reduce steady-state concentrations of BCOD kinase mRNA after a 48 h culture, and this was correlated with a 2-fold increase in the dephosphorylated form of the BCOD complex. The half-life of the kinase mRNA in H4IIE cells was not affected by dexamethasone treatment. Therefore, the decrease in the steady-state kinase mRNA level resulting from dexamethasone treatment was not caused by changes in mRNA stability, which raised the possibility of regulation at the level of gene transcription. To identify the negative glucocorticoid-responsive element in the kinase promoter, nested deletion constucts in the 3.0 kb promoter region were examined in H4IIE cells cultured in the presence or absence of dexamethasone. No significant differences in promoter activity were observed on either transient or stable transfection. The data showed that the glucocorticoid-responsive element was located outside the 3. 0 kb promoter region. At the physiological level, hepatic BCOD kinase mRNA levels were reduced in rats injected intraperitoneally with dexamethasone. This effect was liver-specific, and was not detected in other tissues. These results suggest that the down-regulation of kinase gene expression by glucocorticoids is mediated through a liver-specific or -enriched transcription factor(s).
Huang
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YS
Department of Biochemistry and the Biochemistry and Molecular Biology Graduate Program, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75235, USA.
Chuang
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DT
D000818:Animals; D002478:Cells, Cultured; D003609:Dactinomycin / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology*; D018753:Diet, Protein-Restricted; D015536:Down-Regulation / Q000187:drug effects*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006207:Half-Life; D007438:Introns / Q000235:genetics; D008099:Liver / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D008297:Male; D008930:Mitochondria, Liver / Q000201:enzymology*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011494:Protein Kinases / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D020218:Response Elements / Q000235:genetics; D017384:Sequence Deletion; D014162:Transfection; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D003609:Dactinomycin; D003907:Dexamethasone; D011494:Protein Kinases; C035627:(3-methyl-2-oxobutanoate dehydrogenase (lipoamide)) kinase