Association between hepatocyte nuclear factor 6 (HNF-6) and FoxA2 DNA binding domains stimulates FoxA2 transcriptional activity but inhibits HNF-6 DNA binding.
In previous studies we used transgenic mice or recombinant adenovirus infection to increase hepatic expression of forkhead box A2 (FoxA2, previously called hepatocyte nuclear factor 3beta [HNF-3beta]), which caused diminished hepatocyte glycogen levels and reduced expression of glucose homeostasis genes. Because this diminished expression of FoxA2 target genes was associated with reduced levels of the Cut-Homeodomain HNF-6 transcription factor, we conducted the present study to determine whether there is a functional interaction between HNF-6 and FoxA2. Human hepatoma (HepG2) cotransfection assays demonstrated that HNF-6 synergistically stimulated FoxA2 but not FoxA1 or FoxA3 transcriptional activity, and protein-binding assays showed that this protein interaction required the HNF-6 Cut-Homeodomain and FoxA2 winged-helix DNA binding domains. Furthermore, we show that the HNF-6 Cut-Homeodomain sequences were sufficient to synergistically stimulate FoxA2 transcriptional activation by recruiting the p300/CBP coactivator proteins. This was supported by the fact that FoxA2 transcriptional synergy with HNF-6 was dependent on retention of the HNF-6 Cut domain LXXLL sequence, which mediated recruitment of the p300/CBP proteins. Moreover, cotransfection and DNA binding assays demonstrated that increased FoxA2 levels caused a decrease in HNF-6 transcriptional activation of the glucose transporter 2 (Glut-2) promoter by interfering with the binding of HNF-6 to its target DNA sequence. These data suggest that at a FoxA-specific site, HNF-6 serves as a coactivator protein to enhance FoxA2 transcription, whereas at an HNF-6-specific site, FoxA2 represses HNF-6 transcription by inhibiting HNF-6 DNA binding activity. This is the first reported example of a liver-enriched transcription factor (HNF-6) functioning as a coactivator protein to potentiate the transcriptional activity of another liver factor, FoxA2.
Rausa
Francisco M
FM
Department of Molecular Genetics, College of Medicine, University of Illinois at Chicago, Chicago, Illinois 60607, USA.
Tan
Yongjun
Y
Costa
Robert H
RH
D000256:Adenoviridae / Q000235:genetics; D020816:Amino Acid Motifs; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D050881:E1A-Associated p300 Protein; D005982:Glutathione Transferase / Q000378:metabolism; D051542:Hepatocyte Nuclear Factor 3-beta; D051559:Hepatocyte Nuclear Factor 6; D018398:Homeodomain Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008099:Liver / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D008957:Models, Genetic; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015534:Trans-Activators / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; C496502:FOXA2 protein, human; C496504:Foxa2 protein, rat; D051559:Hepatocyte Nuclear Factor 6; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; C496605:ONECUT1 protein, human; C496607:Onecut1 protein, rat; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D051542:Hepatocyte Nuclear Factor 3-beta; D004247:DNA; D008156:Luciferases; D050881:E1A-Associated p300 Protein; C494362:Ep300 protein, rat; D005982:Glutathione Transferase