title: Disruption of a binding site for hepatocyte nuclear factor 4 results in hemophilia B Leyden.
abstract: Hemophilia B Leyden is an X chromosome-linked bleeding disorder characterized by very low plasma levels of blood coagulation factor IX (fIX) during childhood. After puberty, plasma fIX levels gradually rise to a maximum of 60% of normal, probably under the influence of testosterone. Single point mutations in the fIX promoter region of hemophilia B Leyden patients have been reported at -20, -6, -5, +8 and +13. In addition, one promoter mutation (G----C at -26) has been detected that abolishes fIX expression throughout life (M. Ludwig, personal communication). We examined how one of the hemophilia B Leyden mutations (T----A at -20) and the G----C mutation at -26 interfere with fIX gene transcription. We report that the wild-type promoter of the human fIX gene contains a binding site (at nucleotides -34 to -10) for hepatocyte nuclear factor 4 (HNF-4), a member of the steroid hormone receptor superfamily of transcription factors. The binding of HNF-4 is disrupted by both the T----A mutation at -20 and the G----C mutation at -26. Whereas HNF-4 transactivates the wild-type promoter sequence in liver (HepG2) and non-liver (HeLa) cell types quite well, it transactivates the -20 mutated promoter to only a limited extent and the -26 mutated promoter not at all. These data suggest that HNF-4 is a major factor controlling fIX expression in the normal individual and that its inability to bind efficiently to the -20 T----A and the -26 G----C mutated promoter sequence results in hemophilia. Further, the severity of the hemophilia phenotype appears to be directly related to the degree of disruption of HNF-4 binding and transactivation.
authors.affiliation: Department of Hematology, University Medical Center, Leiden, The Netherlands.
authors.forename: M J
authors.initials: MJ
authors.lastname: Reijnen
authors.forename: F M
authors.initials: FM
authors.lastname: Sladek
authors.forename: R M
authors.initials: RM
authors.lastname: Bertina
authors.forename: P H
authors.initials: PH
authors.lastname: Reitsma
mesh_terms: D001483:Base Sequence; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins / Q000378:metabolism; D005164:Factor IX / Q000235:genetics*; D005786:Gene Expression Regulation; D006367:HeLa Cells; D051557:Hepatocyte Nuclear Factor 4; D006801:Humans; D008969:Molecular Sequence Data; D010750:Phosphoproteins; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection
chemical_list: D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; D051557:Hepatocyte Nuclear Factor 4; C403128:MLX protein, human; D010750:Phosphoproteins; D014157:Transcription Factors; C064567:factor IX Leyden; D005164:Factor IX