Transcriptional repression of the C/EBP-alpha and GLUT4 genes in 3T3-L1 adipocytes by tumor necrosis factor-alpha. Regulations is coordinate and independent of protein synthesis.
We have previously demonstrated the ability of tumor necrosis factor-alpha (TNF) to down-regulate the expression of GLUT4 (insulin-responsive glucose transporter) and C/EBP-alpha (CCAAT/enhancer-binding protein) (Stephens J. M., and Pekala, P. H. (1991) J. Biol. Chem. 266, 21839-21845). As C/EBP-alpha has been suggested to control GLUT4 expression, we have examined the time course for attenuation of transcription of these genes. Run-on transcription assays indicate a coordinate transcriptional repression of both GLUT4 and C/EBP-alpha genes (as well as the 422/aP2 gene, the adipocyte lipid-binding protein, whose expression has also been proposed to be controlled by C/EBP-alpha). Inhibition of transcription was observed within 1 h of TNF addition, with maximal suppression observed after 4 h. The inhibition was not blocked by cycloheximide. Okadaic acid treatment (1 h, 0.5 microM) also resulted in the coordinate transcriptional repression of the C/EBP-alpha, GLUT4, and 422/aP2 genes, consistent with involvement of a kinase-phosphatase system in the regulation of these genes. The decrease in C/EBP-alpha protein content was detectable 4 h after TNF addition and declined to 25% of controls within 24 h. A minor decrease in the protein content of GLUT4 was observed during the first 24 h of exposure to TNF; however, after 72 h of exposure GLUT4 protein was not detectable. The rapid coordinate transcriptional regulation of C/EBP-alpha, GLUT4, and 422/aP2 by TNF in the presence of cycloheximide suggests that the TNF-induced loss of GLUT4 protein may be mediated by a post-translational modification of an existing transcription factor. However, the rapid loss of C/EBP-alpha protein may be a contributing factor to further transcriptional suppression of the GLUT4 gene at the later time points. In addition to the transcriptional effect, we report that TNF-induced destabilization of these mRNAs contributes to decreased expression of all three genes.
Stephens
J M
JM
Department of Biochemistry, School of Medicine, East Carolina University, Greenville, North Carolina 27834.
Pekala
P H
PH
D016475:3T3 Cells; D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000273:Adipose Tissue / Q000166:cytology / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D022762:CCAAT-Enhancer-Binding Proteins; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000235:genetics*; D004988:Ethers, Cyclic / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D051379:Mice; D009004:Monosaccharide Transport Proteins / Q000235:genetics*; D009687:Nuclear Proteins / Q000235:genetics*; D019319:Okadaic Acid; D014158:Transcription, Genetic; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D004988:Ethers, Cyclic; D009004:Monosaccharide Transport Proteins; D009687:Nuclear Proteins; D014409:Tumor Necrosis Factor-alpha; D019319:Okadaic Acid; D015124:8-Bromo Cyclic Adenosine Monophosphate