Regulation of pyruvate dehydrogenase kinase isoform 4 (PDK4) gene expression by glucocorticoids and insulin

Abstract

The pyruvate dehydrogenase complex (PDC) catalyzes the conversion of pyruvate to acetyl-CoA in mitochondria and is a key regulatory enzyme in the oxidation of glucose to acetyl-CoA. Phosphorylation of PDC by the pyruvate dehydrogenase kinases (PDK) inhibits its activity. The expression of the pyruvate dehydrogenase kinase 4 (PDK4) gene is increased in fasting and other conditions associated with the switch from the utilization of glucose to fatty acids as an energy source. Transcription of the PDK4 gene is elevated by glucocorticoids and inhibited by insulin. In this study, we have investigated the factors involved in the regulation of the PDK4 gene by these hormones. Glucocorticoids stimulate PDK4 through two glucocorticoid receptor (GR) binding sites located more than 6,000 base pairs upstream of the transcriptional start site. Insulin inhibits the glucocorticoid induction in part by causing dissociation of the GR from the promoter. Previously, we found that the estrogen related receptor alpha (ERRα) stimulates the expression of PDK4. Here, we determined that one of the ERRα binding sites contributes to the insulin inhibition of PDK4. A binding site for the forkhead transcription factor (FoxO1) is adjacent to the ERRα binding sites. FoxO1 participates in the glucocorticoid induction of PDK4 and the regulation of this gene by insulin. Our data demonstrate that glucocorticoids and insulin each modulate PDK4 gene expression through complex hormone response units that contain multiple factors.

Regulation of Pyruvate Dehydrogenase Kinase 4 (PDK4) by CCAAT/Enhancer-binding Protein β (C/EBPβ)

“…Chromatin Immunoprecipitation Assays-ChIP assays were conducted with modifications following the protocol given by the Millipore Magna ChIP kit (17-610) (42). Rat primary hepatocytes were maintained for 18 h in RPMI 1640 medium containing 5% fetal bovine serum and 5% calf serum.…”

“…After treatment, cross-linking was performed with 1% formaldehyde for 10 min at room temperature, and cells were sonicated as described previously (42). Chromatin preparations were diluted with dilution buffer and protease inhibitor mixture and designated as input samples (no antibody) or immunoprecipitated with the control antibody rabbit IgG (sc-2027, Santa Cruz Biotechnology), anti-C/EBP␤ (sc-150, Santa Cruz Biotechnology), or anti-TR␤ (MA1-216, Thermo Scientific).…”

“…PCR products were analyzed on 2% NuSieve 3:1 agarose (Lonza, Walkersville, MD) and visualized with MultiImage light cabinet with Alphaimager EP software. The following primers were used to amplify portions of the Pdk4 promoter: the proximal Pdk4 promoter (Ϫ591/Ϫ338) FP 5Ј-taaggctatttaggcagttt-3Ј and RP 5Ј-ccagacttgtccttgtttac-3Ј, the TRE (Ϫ1535/Ϫ1228) FP 5Ј-agtgtctccaccagattgt-3Ј and RP 5Ј-ctaagagagctaacctagt-3Ј, and the upstream region (Ϫ6634/ Ϫ6377) FP 5Ј-tatgagaagtgctgcaataa-3Ј and RP 5Ј-atgttaccacaaaccttcat-3Ј (42).…”

Lipid droplet-associated proteins in high-fat fed mice with the effects of voluntary running and diet change

“…The isoform PDK4 is highly expressed in liver, heart and skeletal muscle and is regulated by exercise. Its expression is elevated with diabetes, fasting and other conditions associated with the switch from the utilization of glucose to fatty acids as an energy source [32,33]. It has been suggested that insulin resistance is associated with dysregulation of the PDC in skeletal muscle and that excess insulin would on the other hand down-regulate the expression of PDK4 [25,34,35].…”

An energetic orphan in an endocrine tissue: A revised perspective of the function of estrogen receptor-related receptor alpha in bone and cartilage

“…For instance, a group of genes involved in OXPHOS and known to be downregulated in human diabetic muscle, are direct targets of PGC1a, an action that is dependent on the presence of ERRa. (53) ERRa may also control mitochondrial glucose oxidation to acetyl-coenzyme A (CoA) via the regulation of the pyruvate dehydrogenase kinases 2 and 4 (PDK2-4) in adipocyte and hepatoma cells, respectively, inducing a decrease in the conversion of pyruvate to acetyl-CoA in mitochondria and implicating ERRa as a negative regulator of the oxidation of glucose to acetyl-CoA (54)(55)(56)(57)(58) (Fig. 3).…”