TRANSACTIVATION OF THE ALPHA-1-ACID GLYCOPROTEIN GENE ACUTE-PHASE RESPONSIVE ELEMENT BY MULTIPLE ISOFORMS OF C EBP AND GLUCOCORTICOID RECEPTOR/

Alpha1-acid glycoprotein (AGP) is a major acute phase protein synthesi zed primarily by the liver. The AGP gene is transcriptionally activate d in hepatocytes during the acute phase response to bacterial lipopoly saccharide. In this study, we analyzed an acute phase responsive eleme nt (APRE) located between nucleotide residues -127 to -104 relative to the transcription initiation site of the mouse AGP gene. Binding stud ies show that several trans-acting factors interact with the APRE. Usi ng monospecific antibodies we demonstrate that three isoforms of the C CAAT/enhancer-binding protein (C/EBP) family, namely C/EBPalpha, C/EBP beta, and C/EBPdelta, bind to the APRE. Furthermore, with liver nuclea r protein from control animals, C/EBPalpha is the predominant form tha t binds to the APRE, whereas with nuclear proteins from acute phase-in duced animals, C/EBPalpha is replaced by C/EBPbeta. The mechanism of a ctivation of the AGP gene during the acute phase response appears to i nvolve an exchange of C/EBPalpha by C/EBPbeta. C/EBPdelta does not pla y a role in this reaction. Interestingly, the C/EBP binding site of th e APRE partially overlaps a functional glucocorticoid responsive eleme nt. We present evidence that both purified C/EBPalpha and glucocortico id receptor bind strongly to the APRE. By site-specific mutation, we h ave identified the C/EBP and glucocorticoid receptor binding sites in the APRE. These mutants were used in expression vectors to demonstrate that both C/EBP and glucocorticoid receptor are essential for maximal response to interleukin-6 and dexamethasone. These results demonstrat e that the APRE is a composite binding site for multiple factors that are responsible for the transcriptional control of the mouse AGP. Fina lly, functional analyses indicate that C/EBPalpha, C/EBPbeta, and C/EB Pdelta are strong transcriptional trans-activators of the AGP APRE in hepatoma cells. These data suggest that the regulatory activity of the C/EBP with the APRE in the liver may require interactions with adjace nt proteins.