Role of the CCAAT/enhancer binding protein-alpha transcription factor in the glucocorticoid stimulation of p21waf1/cip1 gene promoter activity in growth-arrested rat hepatoma cells.
The preceding paper (Cha, H. H., Cram, E. J., Wang, E. C., Huang, A. J., Kasler, H. G., and Firestone, G. L. (1998) J. Biol. Chem. 273, 0000-0000(478563) defined a glucocorticoid responsive region within teh promoter of the p21 CDK inhibitor gene that contains a putative DNA-binding site for the transcription factor CCAAT/ enhancer binding protein-alpha (C/EBP alpha). Wild type rat BDS1 hepatoma cells as well as as4 hepatoma cells, which express antisense sequences to C/EBP alpha and ablate its protein production, were utilized to investigate the role of this transcription factor in the glucocorticoid regulation of p21 gene expression. The stimulation of p21 protein levels and promoter activity, as well as inhibition of CDK2-mediated retinoblastoma protein phosphorylation, by the synthetic glucocorticoid, dexamethasone, required the expression of C/EBP alpha. Overexpression of C/EBP alpha in as4 cells rescued the dexamethasone responsiveness of the p21 promoter. Site-directed mutagenesis of the p21 promoter revealed that dexamethasone stimulation of p21 promoter activity required the C/EBP consensus DNA-binding site. Furthermore, in glucocorticoid receptor-defective EDR1 hepatoma cells, dexamethasone failed to stimulate C/EBP alpha and p21 protein expression and promoter activities. Our results have established a functional link between the glucocorticoid receptor signaling pathway that mediates a G1 cell cycle arrest of rat hepatoma cells and the transcriptional control of p21 by a cascade that requires the steroid induction of C/EBP alpha gene expression.
Cram
E J
EJ
Department of Molecular and Cell Biology, University of California, Berkeley 94720, USA.
Ramos
R A
RA
Wang
E C
EC
Cha
H H
HH
Nishio
Y
Y
Firestone
G L
GL
D000818:Animals; D022762:CCAAT-Enhancer-Binding Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000235:genetics*; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000502:physiology*; D003907:Dexamethasone / Q000494:pharmacology; D016193:G1 Phase; D005938:Glucocorticoids / Q000494:pharmacology; D008114:Liver Neoplasms, Experimental / Q000235:genetics* / Q000473:pathology; D016296:Mutagenesis; D009687:Nuclear Proteins / Q000502:physiology*; D011401:Promoter Regions, Genetic; D051381:Rats; D014157:Transcription Factors / Q000502:physiology*
D022762:CCAAT-Enhancer-Binding Proteins; C493958:Cdkn1a protein, rat; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004268:DNA-Binding Proteins; D005938:Glucocorticoids; D009687:Nuclear Proteins; D014157:Transcription Factors; D003907:Dexamethasone; D004247:DNA