Serum Amyloid A Gene Expression under Acute-Phase Conditions Involves Participation of Inducible C/EBP-β and C/EBP-δ and Their Activation by Phosphorylation

Abstract

Serum amyloid A (SAA) is a plasma protein whose synthesis is markedly increased in the liver during the inflammatory process. Previous analysis of SAA promoter function implicated the involvement of the CCAAT/enhancer-binding protein (C/EBP) in controlling this process. In this study, using antibodies against three C/EBP isoforms in DNA-binding and Western blot (immunoblot) assays, we found that in response to inflammatory signals, both C/EBP-8 and C/EBP-β are induced and that their interactions with the SAA promoter element are necessary for the increased SAA gene expression. Cotransfections of liver cells with an SAA promoter-linked reporter chloramphenicol acetyltransferase gene and murine sarcoma virus-expressed C/EBP-δ or C/EBP- β confirm such phenomena. The increased transactivating ability in the presence of the cellular phosphatase inhibitors okadaic acid and sodium orthovanadate, coupled with the observation that dephosphorylation severely inhibits the DNA-binding ability in vitro, implicates a role of phosphorylation in the regulation of the activities of the C/EBP-δ isoform. Consistent with these findings, we have detected higher levels of DNA-binding activity of C/EBP-δ prepared from cells treated with phosphatase inhibitors. We also present evidence that C/EBP-δ is a phosphoprotein. These results suggest that C/EBP-δ is regulated by phosphorylation and, in conjunction with C/EBP-β, is one of the major proteins responsible for the increased transcription of the SAA gene in response to inflammatory stimuli.