Differential regulation of branched-chain alpha-ketoacid dehydrogenase kinase expression by glucocorticoids and acidification in LLC-PK1-GR101 cells.
Acidosis and glucocorticoids (GC) are two catabolic signals associated with chronic renal disease. Previously, we reported that these signals stimulate branched-chain amino acid (BCAA) oxidation in renal tubule cells by increasing both the amount and activation state of branched-chain alpha-ketoacid dehydrogenase (BCKD). Activation of the BCKD complex could result from decreased expression of BCKD kinase, which inhibits BCKD by phosphorylating its E1 alpha subunit. To investigate this possibility, we examined how dexamethasone and acidification (pH 7.0) influence BCKD kinase expression in LLC-PK(1)-GR101 cells. Dexamethasone, a synthetic GC, decreased BCKD kinase protein by 65% (P < 0.05 vs. control), whereas a low pH (i.e., pH 7.0) decreased the amount of kinase by 71% (P < 0.05 vs. control). Either GC or acidification reduced BCKD kinase mRNA by 46% (P < 0.05 vs. control), but the two signals together did not reduce kinase mRNA more than either signal alone. To examine the mechanism(s) leading to lower kinase mRNA, kinase transcription was evaluated by transiently transfecting LLC-PK(1)-GR101 cells with BCKD kinase promoter-luciferase mini-genes containing approximately 3.5 kb of proximal rat kinase promoter. GC, but not acidification, decreased luciferase activity 42% (P < 0.05 vs. control). Nuclear run-on assays confirmed that GC decrease kinase mRNA by attenuating its transcription. Thus two catabolic signals associated with renal failure, GC and acidification, reduce BCKD kinase expression by different mechanisms. These responses lead to an increase in the activation state of BCKD and a resulting acceleration of BCAA degradation.
Wang
Xiaonan
X
Renal Division, Rm. 338 Woodruff Memorial Bldg., 1639 Pierce Drive, Emory University School of Medicine, Atlanta, GA 30322, USA. medrp@emory.edu
Price
S Russ
SR
D002460:Cell Line; D003907:Dexamethasone / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic; D005938:Glucocorticoids / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007684:Kidney Tubules / Q000187:drug effects / Q000201:enzymology*; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism
D005938:Glucocorticoids; D012333:RNA, Messenger; D003907:Dexamethasone; D011494:Protein Kinases; C035627:(3-methyl-2-oxobutanoate dehydrogenase (lipoamide)) kinase