Haemophilia B Leyden: the effect of mutations at position +13 on the liver-specific transcription of the factor IX gene.

Haemophilia B Leyden is characterized by low plasma levels (< or = 1-13% of normal) of blood coagulation factor IX during childhood. After the onset of puberty, plasma factor IX levels gradually rise, probably under the influence of androgens. Single point mutations have been detected in the factor IX promoter at -21, -20, -6, -5, +6, +8 and +13. This paper examines how two of these mutations (a deletion of an A, and an A-->G substitution at +13) interfere with normal factor IX gene transcription. It is shown that both mutations do impair factor IX promoter activity in transiently transfected HepG2 cells. The mutations at +13 lie in a region (+1 to +18) that is considered to contain a binding site for the CCAAT/enhancer binding protein. Transactivation by the CCAAT/enhancer binding protein alpha of the wild-type and mutated factor IX promoter (-192 to +38) resulted in an approximately four-fold and approximately two-fold, respectively, increase of CAT activity. Gel mobility shift assays revealed that the binding of the CCAAT/enhancer binding protein alpha is disrupted by both the deletion of an A, and the A-->G substitution at +13. The role of two additional bZIP factors, D-site binding protein and liver-enriched transcriptional activator protein, in the binding and activation of the +13 factor IX promoter region was examined. The D-site binding protein binds to the factor IX promoter region (+1 to +18) in gel mobility shift assays. The deletion of an A at +13 does not interfere with the binding of the D-site binding protein.(ABSTRACT TRUNCATED AT 250 WORDS)