Cooperative binding of upstream stimulatory factor and hepatic nuclear factor 4 drives the transcription of the human apolipoprotein A-II gene.
The activity of the human apoA-II promoter is controlled by a synergistic interaction of the distal enhancer and the proximal promoter. An important role in apoA-II promoter activity is exerted by a transcription factor, designated CIIIB1, which binds to the proximal element AB and the distal elements of the enhancer, K and L. In the present communication we establish that CIIIB1 corresponds to the previously described factor, upstream stimulatory factor (USF) using the following criteria. (a) Purification of CIIIB1 by affinity chromatography provided a heat-stable protein with an apparent molecular mass of 45 kDa that cross-reacted with anti-USF1 and -USF2a antibodies; (b) CIIIB1 bound to the elements AB, K, and L was supershifted by these antibodies; (c) the heterodimer USF1/2a is the predominant form that corresponds to CIIIB1. Cotransfection experiments in HepG2 cells established the functional significance of USF in apoA-II transcription. It was found that the minimal promoter AB was transactivated by USF2a. In addition, all three E-box motifs present in elements AB, K and L are necessary for maximum transactivation by USF2a. A dominant negative form of USF2a inhibits the activity of apoA-II promoter. The USF1/2a heterodimer, which is naturally expressed in the liver, is as efficient as the USF2a homodimer in the transactivation of apoA-II promoter/enhancer constructs. Cotransfection experiments in COS-1 cells showed that hepatic nuclear factor 4 (HNF-4) synergized with USF2a in the transactivation of the apoA-II promoter. In addition, we showed that HNF-4 and USF2a bind to the enhancer cooperatively. This may account for the transcriptional synergism observed between USF and HNF-4 in the transactivation of the apoA-II promoter.
Ribeiro
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A
U505 INSERM, Université Pierre et Marie Curie, Institut des Cordeliers, 15 rue de l'Ecole de Médecine, 75006 Paris, France.
Pastier
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D
Kardassis
D
D
Chambaz
J
J
Cardot
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P
D000818:Animals; D016633:Apolipoprotein A-II / Q000235:genetics*; D001483:Base Sequence; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D019556:COS Cells; D002460:Cell Line; D002846:Chromatography, Affinity; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins; D018257:Helix-Loop-Helix Motifs; D051557:Hepatocyte Nuclear Factor 4; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D051381:Rats; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D051720:Upstream Stimulatory Factors
D016633:Apolipoprotein A-II; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; D051557:Hepatocyte Nuclear Factor 4; C403128:MLX protein, human; D009687:Nuclear Proteins; D010750:Phosphoproteins; D014157:Transcription Factors; C497181:USF1 protein, human; D051720:Upstream Stimulatory Factors; C497182:Usf1 protein, rat; C068859:nuclear factor CIIIB1; D004247:DNA