Hormonal Regulation of the Phosphoenolpyruvate Carboxykinase Gene

• Published in: The Journal of biological chemistry 2000-02, Vol.275 (8), p.5804-5809
• Main Authors: Crosson, Sean M., Roesler, William J.
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.275.8.5804

The CCAAT/enhancer-binding protein α (C/EBP) is a transcription factor that trans-activates a number of metabolically important genes. Previous work has demonstrated that C/EBPα and C/EBPβ have the potential to mediate the cAMP responsiveness of phosphoenolpyruvate carboxykinase (PEPCK) in liver cells. However, these studies used GAL4 fusion proteins and artificial promoter-reporter gene vectors in transfection experiments; as a result, these studies only indicated that both isoforms had the potential to mediate the hormonal response and not which isoform actually participated in vivo. To address this issue, we produced hepatoma cell lines that stably expressed either a dominant negative inhibitor or antisense RNA for these two main liver C/EBP isoforms. Inhibition of all C/EBP isoforms via expression of the dominant negative protein eliminated cAMP responsiveness, and reduced glucocorticoid responsiveness, of the endogenous PEPCK gene in hepatoma cells. Antisense directed against C/EBPα mRNA, which reduced C/EBPα protein levels by nearly 80%, also significantly reduced the cAMP responsiveness of the endogenous PEPCK promoter, whereas antisense directed against C/EBPβ was without effect. There was no major alteration in cAMP signaling in the C/EBPα antisense cells, as cAMP induction of the C/EBPβ gene was similar to that in wild-type H4IIE cells. These data suggest that the α-isoform of C/EBP is specifically utilized for mediating the cAMP responsiveness of the PEPCK gene.