Functional analysis of developmentally regulated chromatin-hypersensitive domains carrying the alpha 1-fetoprotein gene promoter and the albumin/alpha 1-fetoprotein intergenic enhancer.
During liver development, the tandem alpha 1-fetoprotein (AFP)/albumin locus is triggered at the AFP end and then asymmetrically enhanced; this is followed by autonomous repression of the AFP-encoding gene. To understand this regulation better, we characterized the two early developmental stage-specific DNase I-hypersensitive (DH) sites so far identified in rat liver AFP/albumin chromatin: an intergenic DH-enhancer site and the AFP DH-promoter site. Mutation-transfection analyses circumscribed the DH-enhancer domain to a 200-bp DNA segment stringently conserved among species. Targeted mutations, DNA-protein-binding assays, and coexpression experiments pinpointed C/EBP as the major activatory component of the intergenic enhancer. Structure-function relationships at the AFP DH-promoter site defined a discrete glucocorticoid-regulated domain activated cooperatively by HNF1 and a highly specific AFP transcription factor, FTF, which binds to a steroid receptor recognition motif. The HNF1/FTF/DNA complex is deactivated by glucocorticoid receptors or by the ubiquitous factor NF1, which eliminates HNF1 by competition at an overlapping, high-affinity binding site. We propose that the HNF1-NF1 site might serve as a developmental switch to direct autonomous AFP gene repression in late liver development. We also conclude that the intergenic enhancer is driven by C/EBP alpha primarily to fulfill albumin gene activation functions at early developmental stages. Factor FTF seems to be the key regulator of AFP gene-specific functions in carcinoembryonic states.
Bernier
D
D
Le Centre de recherche en cancérologie, Université Laval, L'Hôtel-Dieu de Québec, Canada.
Thomassin
H
H
Allard
D
D
Guertin
M
M
Hamel
D
D
Blaquière
M
M
Beauchemin
M
M
LaRue
H
H
Estable-Puig
M
M
Bélanger
L
L
D000418:Albumins / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D002843:Chromatin / Q000378:metabolism*; D002845:Chromatography; D002874:Chromosome Mapping; D004247:DNA / Q000378:metabolism; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins; D003850:Deoxyribonuclease I / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D051537:Hepatocyte Nuclear Factor 1; D051538:Hepatocyte Nuclear Factor 1-alpha; D051539:Hepatocyte Nuclear Factor 1-beta; D008099:Liver / Q000502:physiology; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000378:metabolism; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D000509:alpha-Fetoproteins / Q000235:genetics*
D000418:Albumins; D002843:Chromatin; D004268:DNA-Binding Proteins; D051538:Hepatocyte Nuclear Factor 1-alpha; C496489:Hnf1a protein, rat; D009687:Nuclear Proteins; D011965:Receptors, Glucocorticoid; D014157:Transcription Factors; D000509:alpha-Fetoproteins; D051537:Hepatocyte Nuclear Factor 1; D051539:Hepatocyte Nuclear Factor 1-beta; D004247:DNA; D003850:Deoxyribonuclease I