Bilirubin UDP-glucuronosyltransferase Gene Research Articles

The Relationship Between Serum Bilirubin and Crohn’s Disease

The oxidative stress is thought to play an important role in Crohn's disease (CD). As serum bilirubin represents the major endogenous antioxidant, this article aimed to evaluate in a clinical study, whether serum bilirubin levels and genes affecting its systemic concentrations are associated with CD. This exploratory case-control study was based on pediatric (n = 119) and adult (n = 504) patients with CD and 370 appropriate healthy control subjects. The (GT)n and (TA)n dinucleotide variations in heme oxygenase 1 (HMOX1) and bilirubin UDP-glucuronosyl transferase (UGT1A1) gene promoters were determined by fragment analysis. Serum bilirubin levels were compared in a subset of 90 cases and 229 controls, for whom biochemical data were available. Substantially lower serum bilirubin levels were detected in patients with CD compared with controls (7.4 versus 12.1 μmol/L, P < 10). Serum bilirubin levels were significantly lower in patients with CD within all UGT1A1*28 genotypes (P < 0.05). UGT1A1*28 homozygotes with wild-type NOD2 gene variant exhibited significant delay in CD manifestation (P = 0.004), while the protective effect of UGT1A1*28 homozygosity was lost in those patients with mutated NOD2 gene. No associations between CD risk and individual HMOX1 gene variants were observed. CD is associated with significantly low serum bilirubin levels, most likely as a result of increased oxidative stress accompanying this inflammatory disease. UGT1A1*28 allele homozygosity, responsible for higher bilirubin levels, seems to be an important modifier of CD manifestation.

Analysis of bilirubin UDP-glucuronosyltransferase gene mutations in an unusual Crigler-Najjar syndrome patient

Crigler-Najjar (CN) syndrome is a rare autosomal recessive inherited disorder characterized by non-hemolytic, unconjugated hyperbilirubinemia. The levels of serum bilirubin and the response to phenobarbital treatment have been used to classify CN syndrome into two types: CN I and II. Mutations of the UGT1A1 gene have been found to be responsible for cases of CN syndrome. In the present study, the clinical features of a boy with an unusual type of CN syndrome were analysed. A DNA sample was obtained from the patient, and the promoter region, the exons and flanking intronic sequences of the UGT1A1 gene were analysed using the polymerase chain reaction and sequencing. The case was similar to CN typeI in clinical features, but the therapeutic efficacy in the patient was superior to that typically observed in CN typeII disease. Sequencing revealed compound heterozygous mutations, c.211G>A (p.G71R), c.1470C>T (p.D490D) and a normal homozygous A[TA]6TAA. No similar case has been reported worldwide and, considering the specific clinical features and therapeutic efficacy, a distinct type of CN was suspected. The phenotype of this unusual CN syndrome patient may be associated with the specific genotype.

Cord blood bilirubin level in relation to bilirubin UDP‐glucuronosyltransferase gene missense allele in Chinese neonates

To investigate bilirubin UDP-glucuronosyltransferase (UGT1A1) gene allele in healthy Chinese neonates, their cord bilirubin level and the subsequent hyperbilirubinemia to determine relationships among them. Cord blood of 48 neonates was obtained to determine the exon 1 of UGT1A1 gene, total serum bilirubin, albumin, glutamic-pyruvic transaminase (GPT), glutamic-oxalacetic transaminase (GOT) and haemoglobin (Hb) concentration. Neonatal jaundice was assessed by measurement of transcutaneous bilirubin (TCB) and serum bilirubin. Neonates were divided into two groups according to mutant or normal allele to compare the variables. Nineteen infants had the nucleotide 211 G-->A allele, 3 had the heterozygous variation (686C-->A, 845 A-->T, 231G-->A). In the 211 A allele group, cord bilirubin was significantly higher than in the 211 G allele group (p = 0.034), but there were no differences in albumin (p = 0.678), GPT (p = 0.460), GOT (p = 0.440) and Hb (p = 0.886). The TCB (at 48, 96 h), the frequency of the hyperbilirubinemia and prolonged jaundice were also significantly higher in the 211 A allele group. The UGT1A1 gene codon G71R allele is a risk factor for neonatal hyperbilirubinemia in the Chinese population. Its effect on bilirubin metabolism may present early on, as well as late in foetal life.

UGT1A1(TA)n promoter polymorphism—A new case of a (TA)8 allele in Caucasians

Gilbert's syndrome is a mild hereditary unconjugated hyperbilirubinemia caused by mutations in the bilirubin UDP-glucuronosyltransferase gene (UGT1A1). The causative mutation in Caucasians is almost exclusively a TA dinucleotide insertion in the TATA box of the UGT1A1 promoter. Affected individuals are homozygous for the variant promoter and have 7 instead of 6 TA repeats. The aim of the present study was to determine the genotypes of UGT1A1(TA)n promoter polymorphism in the healthy Slovenian population and to investigate the association of genotypes with serum bilirubin levels. 236 healthy subjects were genotyped by single-strand conformation polymorphism analysis, which was validated by sequence analysis. The frequencies of genotypes were as follows: (TA)6/6 (38.1%), (TA)6/7 (47.9%), (TA)7/7 (13.6%). There was a statistically significant association of genotypes with serum bilirubin levels (p<0.001). Subjects with genotype (TA)7/7 had the highest and subjects with genotype (TA)6/6 the lowest total serum bilirubin levels. One individual in the group had the rare genotype (TA)7/8 (0.4%). Analysis of his family showed the following genotypes: (TA)6/8 in his father and sister and (TA)7/8 in his two brothers. In conclusion, the frequency of UGT1A1(TA)n promoter polymorphism genotypes was determined for the first time in the Slovenian population and is similar to frequencies observed in other Caucasian populations. The extremely rare (TA)8 allele in Caucasians was found also in Slovenians.

Hematologically important mutations: Bilirubin UDP-glucuronosyltransferase gene mutations in Gilbert and Crigler–Najjar syndromes

Gilbert and Crigler-Najjar syndromes are familial unconjugated hyperbilirubinemias caused by genetic lesions involving a single complex locus encoding for bilirubin UDP-glucuronosyltransferase (UGT1A1) gene. Over the last years a number of different mutations affecting this gene have been characterized. In this report is provided a summary of reported Gilbert and Crigler-Najjar syndromes associated UGT1A1 gene mutations.

Immune response to lentiviral bilirubin UDP-glucuronosyltransferase gene transfer in fetal and neonatal rats

Gene therapy for inherited disorders might cause an immune response to the therapeutic protein. A solution would be to introduce the gene in the fetal or neonatal period, which should lead to tolerization. Lentiviral vectors mediate long-term gene expression, and are well suited for gene therapy early in development. A model for fetal or neonatal gene therapy is the inherited disorder of bilirubin metabolism, Crigler-Najjar disease (CN). The absence of bilirubin UDP-glucoronyltransferase (UGT1A1) activity in CN patients causes high serum levels of unconjugated bilirubin and brain damage in infancy. CN is attractive for the development of gene therapy because the mutant Gunn rat closely mimics the human disease. Injection of UGT1A1 lentiviral vectors corrected the hyperbilirubinemia for more than a year in rats injected as fetuses and for up to 18 weeks in rats injected the day of birth. UGT1A1 gene transfer was confirmed by the presence of bilirubin glucuronides in bile. All animals injected with UGT1A1 lentiviral vectors developed antibodies to UGT1A1. Animals injected with green fluorescent protein (GFP) lentiviral vectors did not develop antibodies to GFP. Our results indicate that fetal and neonatal gene therapy with immunogenic proteins such as UGT1A1 does not necessarily lead to tolerization.

The Polymorphism c.-3279T&amp;gt;G in the Phenobarbital-Responsive Enhancer Module of the Bilirubin UDP-Glucuronosyltransferase Gene Is Associated with Gilbert Syndrome

Hepatic glucuronization of water-insoluble bilirubin is catalyzed by isoenzyme 1A1 of UDP-glucuronosyltransferase (UGT1A1), which is essential for efficient biliary excretion of bilirubin. Mild hyperbilirubinemia, usually <50 μmol/L, is associated with Gilbert syndrome (GS) and is thought to reflect a small reduction in UGT1A1 activity (1)(2). The main cause of GS in all studied populations is a TA insertion in the repetitive TATA box of the gene promoter(3)(4)(5)(6), which usually consists of 6 repeats. This (TA)7 allele is extremely common, occurring with an estimated frequency of 38.7% in the white population and 16% in the Asian population(7). In persons with African ancestry, the number of alleles carrying 7 or more repeats is reported to be 49.5%(7). On the basis of the frequencies observed in Caucasians in several studies of smaller series, Beutler et al.(7) calculated a homozygosity frequency of 15%. This predicted value is higher than that in patients with a clinical diagnosis of GS, suggesting incomplete penetrance of the (TA)7 allele. Moreover, the fact that some GS patients are neither homozygous for the (TA)7 allele nor have other UGT1A1 genetic deficits points toward the existence of other inherited or acquired factors affecting bilirubin metabolism. Phenobarbital treatment for hyperbilirubinemia increases UGT1A1 activity in the liver via a phenobarbital-responsive enhancer sequence with 3 potential nuclear receptor (NR) motifs separated by ∼90 bases (8). From the 5′end, these motifs are named NR4, gtNR1, and NR3. Alterations in this region could be related to hyperbilirubinemia(8). A polymorphism (c.-3279T>G) located in the NR3 motif is associated with a decrease in transcriptional activity, to …

Two linked polymorphic mutations (A(TA)7TAA and T-3279G) of UGT1A1 as the principal cause of Gilbert syndrome.

Gilbert syndrome is a mild hereditary unconjugated hyperbilirubinemia caused by mutations in the bilirubin UDP-glucuronosyltransferase gene (UGT1A1). The mutation, A(TA)7TAA, is thought to be the sole cause of the syndrome in Caucasians, but an enhancer polymorphism (T-3279G) that lowers transcriptional activity has recently been reported. We have tested the linkage of the two mutations in 11 Caucasians and 12 Japanese patients who were homozygous for A(TA)7TAA. All 23 patients were also homozygous for T-3279G, indicating that T-3279G and A(TA)7TAA were linked. The decrease in transcription caused by both mutations together may be essential to the syndrome.

Polymorphisms of heme oxygenase-1 and bilirubin UDP-glucuronosyltransferase genes are not associated with Kawasaki disease susceptibility.

Kawasaki disease (KD) is a systemic vasculitis and occurs among Japanese children at a high incidence. Serum bilirubin and heme oxygenase-1 (HO-1) expression are known to play a significant role in the protection of vascular endothelial cells. Japanese have unique polymorphic distribution patterns of (TA)7 or G71R of the bilirubin UDP-glucuronosyltransferase (B-UGT) gene and of (GT)n repeats of the HO-1 gene. We investigated the relationship of KD susceptibility with these polymorphisms. There were no significant differences in the distribution of allele frequencies and genotypes of these polymorphisms between KD patients and controls. These polymorphisms are not associated with KD susceptibility.

TA-insertion mutation in bilirubin-UDP glucuronosyltransferase gene (UGT1A1) promoter in Indian patients with Gilbert's syndrome

TA-insertion mutation in bilirubin-UDP glucuronosyltransferase gene (UGT1A1) promoter in Indian patients with Gilbert's syndrome

Correlation of mutational analysis to clinical features in Taiwanese patients with Gilbert’s syndrome

OBJECTIVES: Mutations in the promoter as well as in the coding region of the bilirubin UDP-glucuronosyltransferase gene (UGT1A1) have been found to be associated with Gilbert’s syndrome. However, the genetic basis of Gilbert’s syndrome in our population and correlation of these mutations to fasting serum bilirubin levels in patients with Gilbert’s syndrome remain to be clarified. METHODS: We applied polymerase chain reaction–based direct-sequencing assays to examine mutations in UGT1A1 gene in 20 unrelated Gilbert’s patients and in a family with Gilbert’s syndrome. RESULTS: We studied three mutations that were previously reported to be associated with Gilbert’s syndrome ( i.e., the TATAA-box mutation, Gly71Arg, and Pro229Gln) in 20 patients with Gilbert’s syndrome. Of the patients, 16, five, and six were found to have the TATAA-box, Gly71Arg and Pro229Gln mutations, respectively. Seven patients had simultaneous mutations both in the TATAA box and in the coding region. Of note, all six patients with Pro229Gln also had the TATAA-box mutation. Localization of Pro229Gln on the allele containing the TATAA-box mutation was demonstrated in a family with Gilbert’s syndrome. The patients simultaneously heterozygous for both the TATAA-box mutation and Gly71Arg usually had serum bilirubin levels similar to those found in the patients homozygous for the TATAA-box mutation, but usually higher than those found in the patients heterozygous for the TATAA-box mutation alone. On the other hand, concurrence of Pro229Gln in patients with TATAA-box mutation or with Gly71Arg did not significantly affect serum bilirubin levels. CONCLUSIONS: The TATAA-box mutation and Gly71Arg are the major causes for Gilbert’s syndrome in our population. Concurrence of mutations of Gly71Arg and TATAA-box usually exerts a synergistic effect on hyperbilirubinemia. Pro229Gln, which is regularly linked to the TATAA-box mutation, may not have a significant effect on serum bilirubin levels.

Correlation of mutational analysis to clinical features in Taiwanese patients with Gilbert's syndrome.

Mutations in the promoter as well as in the coding region of the bilirubin UDP-glucuronosyltransferase gene (UGT1A1) have been found to be associated with Gilbert's syndrome. However, the genetic basis of Gilbert's syndrome in our population and correlation of these mutations to fasting serum bilirubin levels in patients with Gilbert's syndrome remain to be clarified. We applied polymerase chain reaction-based direct-sequencing assays to examine mutations in UGT1A1 gene in 20 unrelated Gilbert's patients and in a family with Gilbert's syndrome. We studied three mutations that were previously reported to be associated with Gilbert's syndrome (i.e., the TATAA-box mutation, Gly71Arg, and Pro229Gln) in 20 patients with Gilbert's syndrome. Of the patients, 16, five, and six were found to have the TATAA-box, Gly71Arg and Pro229Gln mutations, respectively. Seven patients had simultaneous mutations both in the TATAA box and in the coding region. Of note, all six patients with Pro229Gln also had the TATAA-box mutation. Localization of Pro229Gln on the allele containing the TATAA-box mutation was demonstrated in a family with Gilbert's syndrome. The patients simultaneously heterozygous for both the TATAA-box mutation and Gly71Arg usually had serum bilirubin levels similar to those found in the patients homozygous for the TATAA-box mutation, but usually higher than those found in the patients heterozygous for the TATAA-box mutation alone. On the other hand, concurrence of Pro229Gln in patients with TATAA-box mutation or with Gly71Arg did not significantly affect serum bilirubin levels. The TATAA-box mutation and Gly71Arg are the major causes for Gilbert's syndrome in our population. Concurrence of mutations of Gly71Arg and TATAA-box usually exerts a synergistic effect on hyperbilirubinemia. Pro229Gln, which is regularly linked to the TATAA-box mutation, may not have a significant effect on serum bilirubin levels.

GILBERT SYNDROME ASSOCIATED WITH β-THALASSEMIA

The authors investigated whether the considerable variability in serum bilirubin levels (STB) found in transfusion-dependent g -thalassemia, g -thal intermedia, and heterozygous g -thalassemia individuals could be related to the coexistence of Gilbert syndrome (GS). The promoter region [A(TA) n TAA] of the bilirubin UDP-glucuronosyltransferase gene (UGT1A1) was analyzed in a total of 128 g -thalassemia individuals (108 transfusion-dependent g -thal patients, 20 very mild g -thal intermedia) and in 33 g -thal heterozygotes. The control group consisted of 70 healthy children with no history of anemia. The frequency of GS genotype (TA) 7 /(TA) 7 did not differ significantly between the groups studied. A significant difference was observed between serum bilirubin levels (STB) and GS genotypes (TA) 7 /(TA) 7 and (TA) 6 /(TA) 7 and also between (TA) 7 /(TA) 7 and (TA) 6 /(TA) 6 for all groups examined. These results confirm that the (TA) 7 /(TA) 7 GS genotype is one of the factors accounting for the hyperbilirubinemia observed in g -thalassemia major, intermedia, and heterozygous individuals.

Expression of bilirubin UDP-glucuronosyltransferase (bUGT) throughout fetal development: intrasplenic transplantation into Gunn rats to correct enzymatic deficiency.

The aim of this work was to determine the pattern of expression of hepatic bilirubin UDP-glucuronosyltransferase throughout fetal development in rats, with the purpose of using fetal hepatocytes at the most appropiate stage of development for transplantation into Gunn rats lacking bilirubin UDP-glucuronosyltransferase activity and then assessing the therapeutic capacity of the implants. The results show that at day 13 of gestational life there is already bilirubin UDP-glucuronosyltransferase gene expression. Twenty-one-day fetal hepatocyte transplantation was also performed into the spleens of hyperbilirubinemic Gunn rats, when alpha-fetoprotein mRNA is still detectable. At 15, 30, and 90 days after transplantation, a mild decrease in total bilirubin serum levels was observed. An increase in bile conjugated bilirubin also was observed at 30 and 90 days. These data suggest the favorable evolution of transplanted cells and show its feasibility for therapy.

Functional Analysis of a Mutation Occurring between the Two In-frame AUG Codons of Human Angiotensinogen

Angiotensinogen (ANG) is the specific substrate of the renin-angiotensin system, a major participant in blood pressure control. We have identified a natural mutation at the -30 amino acid position of the angiotensinogen signal peptide, in which an arginine is replaced by a proline (R-30P). Heterozygous individuals with R-30P showed a tendency to lowered plasma angiotensinogen level (1563 ng of ANG I/ml (range 1129-1941)) compared with normal individuals in the family (1892 ng of ANG I/ml (range 1603-2072)). Human angiotensinogen mRNA has two in-phase translation initiation codons (AUG) starting upstream 39 and 66 nucleotides from the cap site. R-30P occurs in a cluster of basic residues adjacent to the first AUG codon that may affect intracellular sorting of the nascent protein. Pulse-chase experiments in transiently transfected cultured cells revealed that the R-30P mutation was associated with reduced amounts of both intra- and extracellular protein. In a cell-free system, we found that two forms of native angiotensinogen were generated by alternative initiation of translation at either AUG codon. Alteration of either the first or second AUG codons abolished the synthesis of the longer and the shorter form of native angiotensinogen, respectively. Furthermore, the rate of secretion of the shorter form was lower than that of the longer form. By transplanting angiotensinogen signal peptide onto green fluorescence protein, however, we found that both forms of the signal peptide could target green fluorescence protein, normally localized in the cytoplasm, to the secretory pathway. Although the R-30P mutation may not affect intracellular sorting of angiotensinogen in a qualitative manner, it leads to a quantitative reduction in the net secretion of mature angiotensinogen through decreased translocation or increased residence time in the endoplasmic reticulum.

Intermittent jaundice in patients with acute leukaemia: a common mutation of the bilirubin uridine-diphosphate glucuronosyltransferase gene among Asians.

The Gly71Arg mutation of the hepatic bilirubin UDP glucuronosyl-transferase (B-UGT) gene associated with Gilbert syndrome prevails among Japanese and its gene frequency is 0.13. Among 20 patients with acute leukaemia, 4 patients showed intermittent unconjugated hyperbilirubinaemia during the course of combined chemotherapy. The Gly71Arg mutation was detected in all 4 patients with hyperbilirubinaemia, but was not found in 16 patients without hyperbilirubinaemia. Two of them were heterozygotes and one was a homozygote for the Gly71Arg mutation, and the other was a compound heterozygote of the Gly71Arg mutation and TA insertion mutation in the TATA box of the B-UGT gene. In addition to the complications leading to hyperbilirubinaemia, including liver damage due to drugs, viral infections or tumour cell infiltrations and alloimmune haemolysis, carrier status for the Gly71Arg mutation should be considered in a patient with leukaemia showing intermittent hyperbilirubinaemia during the course of chemotherapy, especially among Japanese, Koreans and Chinese owing to its prevalence in those populations.

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 K m (2.5 μM) 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.

Gilbert syndrome caused by a homozygousmissense mutation (Tyr486Asp) of bilirubinUDP-glucuronosyltransferase gene

We report a case of Gilbert syndrome caused by a homozygous missense mutation (Tyr486Asp) of the bilirubin UDP-glucuronosyltransferase gene. Homozygous missense mutations of the gene have previously been recognized as responsible for Crigler-Najjar syndrome type II. We conclude that Gilbert syndrome in some patients results from homozygous missense mutations of the UDP-glucuronosyltransferase gene.

Analysis of the promoter of human bilirubin UDP-glucuronosyltransferase gene (UGT1*1) in relevance to Gilbert's syndrome

Bilirubin UDP-glucuronosyltransferase 1 (B-UGT 1) is an enzyme responsible for hepatic bilirubin glucuronidation and in Gilbert's syndrome, its activity is decreased to 30% of normal. About one third of the patients we analyzed with the syndrome had a homozygous TATA box mutation in UGT1*1 coding for B-UGT 1, i.e. (TA) 7TAA instead of (TA) 6TAA. Since the frequency was much higher than in the normal Japanese population (≈1%), the mutation was considered to be closely associated with the etiology of the syndrome. Since these patients did not have any structural mutations in UGT1*1, we suspected a transcriptional defect of the gene. We analyzed the promoter region (up to −3190) of UGT1*1 by transient transfection assay to identify transcriptionally regulatory sequences and found two elements; one was between −1362 and −1220 and the other between −113 and −70. The proximal one (PE) consisted of two elements, E-box (−104 to −95) and an HNF-1 site (−91 to −79). The two TATA boxes were compared in promoter activity, but no substantial difference was observed. Distal element (DE) and PE were then analyzed in these patients' DNA ( n=6) by PCR and direct sequencing. The same homozygous mutation (C-1353A) was found in two of them, but the reduction in promoter activity was only 15%. These results suggest that the TATA box mutation itself is not the major cause of the syndrome and may be genetically linked to an, as yet, unidentified defect in the further upstream or in the intronic region of UGT1*1.

Hyperbilirubinaemia in heterozygous beta-thalassaemia is related to co-inherited Gilbert's syndrome.

The reasons why heterozygotes for beta-thalassaemia have considerable variation in serum bilirubin levels are unknown. High levels of bilirubin could be related to the co-inherited Gilbert's syndrome, determined either by mutations of the coding region or by variation in the A(TA)nTAA motif of the promoter of the bilirubin UDP-glucuronosyltransferase gene (UGT-1). We sequenced the coding and the promoter region of UGT-1A or characterized the A(TA)nTAA motif of the promoter by denaturing gel electrophoresis of radioactive amplified products. The results were correlated with bilirubin levels in 49 beta-thalassaemia heterozygotes for codon 39 (CAG --> TAG) nonsense mutation. 21 normal individuals and 32 unrelated patients with Gilbert's syndrome served as controls. The coding sequence region of the UGT-1A was normal. Five beta-thalassaemia heterozygotes, who were homozygous for the extra (TA) bases in the A(TA)nTAA element of the promoter of UGT-1A, the configuration present in homozygosity in Gilbert's syndrome, had higher bilirubin levels compared to those with the (TA)6/(TA)7 or (TA)6/(TA)6 configurations. In the group of 32 patients with Gilbert's syndrome, 31 of whom had the (TA)7/(TA)7 configuration, we detected 14 heterozygotes for beta-thalassaemia, a figure much higher than predicted on the basis of the carrier rate. Homozygosity for the (TA)7 motif, the typical promoter configuration of Gilbert's syndrome, is one of the factors determining hyperbilirubinaemia in heterozygous beta-thalassaemia.