Participation of two isoforms of C/EBPbeta transcription factor in the acute-phase regulation of the rat haptoglobin gene.
Previous analyses of the mechanism of the transcriptional induction of the rat haptoglobin (Hp) gene during acute-phase (AP)-reaction have revealed the involvement of several trans-acting nucleoproteins (NPs) in controlling this process. In this study, by using antibodies against C/EBPbeta factor in Western immunoblot assay, we found that rat liver trans-acting NPs p35 and p20 are two characteristic C/EBPbeta isoforms whose expression is induced under AP-conditions. DNA-binding assays identified the binding sites for these two C/EBPbeta proteins in the functionally defined elements A and C of the rat Hp gene and also revealed that they have specific binding affinity towards these elements. Under non-induced conditions, p35 was the only C/EBPbeta binding factor; however, upon AP-conditions both, 35 kDa- and 20 kDa-C/EBPbeta binding activities were significantly induced suggesting that these interactions are necessary for the activation of the Hp gene. By in vitro phosphorylation assay and selective proteolysis, we also present evidence that p35 requires phosphorylation for its DNA binding ability. Thus, we conclude that increase in binding of C/EBPbeta isoforms during AP-reaction occurs through their upregulation and structural modification.
Grigorov
I
I
Molecular Biology Laboratory, Institute for Biological Research, 29 November 142, Belgrade, 11060, Yugoslav Federal Republic.
Milosavljević
T
T
Cvetković
I
I
Petrović
M
M
D000210:Acute-Phase Reaction / Q000235:genetics* / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D022762:CCAAT-Enhancer-Binding Proteins; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006242:Haptoglobins / Q000235:genetics*; D008099:Liver / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D051381:Rats; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D015854:Up-Regulation
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D006242:Haptoglobins; D009687:Nuclear Proteins; D020033:Protein Isoforms; D014157:Transcription Factors; D004247:DNA