Coding defect and a TATA box mutation at the bilirubin UDP-glucuronosyltransferase gene cause Crigler-Najjar type I disease.

Mutations at the bilirubin UDP-glucuronosyltransferase (transferase) gene in a severely hyperbilirubinemic Crigler-Najjar (CN) type I individual was compared with that in a moderately hyperbilirubinemic CN II individual. The CN-I (CF) patient in this study sustained a TATA box insertional mutation which was paired with a coding defect at the second allele, unlike all coding defects previously seen in CN-I patients. The sequence of the mutant TATA box, [A(TA)8A], also seen in the CN-II patient, was compared with that at the wild-type box, [A(TA)7A]. Transcriptional activity with [A(TA)8A] was 10-15% that with the wild-type box when present in the -1.7 kb upstream regulatory region (URR) of the bilirubin transferase UGT1A1 gene which was fused to the chloramphenicol acetyl transferase reporter gene, pCAT 1.7H, and transfected into HepG2 cells. Also, a construct with a TA deletion, [A(TA)6A], was prepared and used as a control; transcriptional activity was 65% normal. The coding region defect, R336W, seen in CF (CN-I) was placed in the bilirubin transferase UGT1A1 [HUG-Br1] cDNA, and its corresponding protein was designated UGT1A1*32. The UGT1A1*32 protein supported 0-10% normal bilirubin glucuronidation when expressed in COS-1 cells. The I294T coding defect seen at the second allele in SM (CN-II) generated the UGT1A1*33 mutant protein which supported 40-55% normal activity with a normal Km (2.5 microM) for bilirubin. The hyperbilirubinemia seen in SM decreased in response to phenobarbital treatment, unlike that seen in CF. Parents of the patients were carriers of the respective mutations uncovered in the offspring. The TATA box mutation paired with a deleterious missense mutation is, therefore, completely repressive in the CN-I patient, and is responsible for a lethal genotype/phenotype; but when homozygous, i.e. paired with itself, as previously reported in the literature, it is far less repressive and generates the mild Gilbert's phenotype.