Identification of two single base substitutions in the UGT1 gene locus which abolish bilirubin uridine diphosphate glucuronosyltransferase activity in vitro.

Abstract

Accumulating evidence indicates that mutations in the human UGTJ gene locus abolish hepatic bilirubin UDP-glucuronosyltransferase activity and cause the subsequent accumulation of bilirubin to toxic levels in patients with Crigler-Najjar type 1 (CN-I). Genetic and biochemical criteria are required to link CN-I with mutations in UGTI. Here we present analysis of mutations at the UGTJ locus in three individuals that were clinically diagnosed with CN-I (two related and one unrelated). Each patient carries a single base substitution that alters conserved residues in the transferase enzyme molecule, serine to phenylalanine at codon 376 and glycine to glutamic acid at codon 309. Each was homozygous for the defect as demonstrated by sequencing and RFLPs. Mutant cDNAs, constructed by site-directed mutagenesis, inserted into expression vectors, and transfected into COS-1 cells, supported the synthesis of the bilirubin transferase protein but only cells transfected with the wild-type cDNA expressed bilirubin UDP-glucuronosyltransferase activity. The data provide conclusive evidence that alterations at Gly 309 and Ser 376 are the genetic basis for CN-I in these families. These results suggest that the two codons, located in conserved regions of the molecule, are part of the active site of the bilirubin enzyme. (J. Clin. Invest. 1994. 93:564-570.)

Impact of UGT1A1 gene variants on total bilirubin levels in Gilbert syndrome patients and in healthy subjects

“…Variants in the promoter and coding regions of UGT1A1 may coexist in GS patients and healthy controls [6,7,9,28,[35][36][37][38]. By direct sequencing of these regions, we identified for the first time seven variants, three in GS patients (c.538G>C; c.1211T>C; c.1423C>T) and four in controls (c.643A>G; c.814A>G; c.1156G>A; c.1475T>C).…”

“…Seven of these variants were new c.538G>C, c.643A>G, c.814A>G, c.1156G>A; c.1211T>C; c.1423C>T; c.1475T>C (Fig. 1), and two were already described in literature, c.674T>G [6] and c.923G>A [28]. Moreover, three of these new variants were detected in GS patients (c.538G>C; c.1211T>C; c.1423C>T) and four in controls (c.643A>G; c.814A>G; c.1156G>A; c.1475T>C).…”

“…The variant c.1423 C > T (p.R475C), also located at donor-binding site, might alter the tertiary structure of the protein. The variant c.923 G > A (p.G308E), initially described by Erps and co-workers [28] is located in a highly conserved residue, and results in lack of enzyme activity. This substitution was described as the second most frequent variant of UGT1A1 gene associated to GS in the Portuguese population [20].…”

De Novo Donor-Specific HLA Antibody Formation in Two Patients With Crigler-Najjar Syndrome Type I Following Human Hepatocyte Transplantation With Partial Hepatectomy Preconditioning

“…First, he presented on day 7 with mycophenolate intoxication. Mycophenolate is primarily metabolized by glucuronidation by UGT1A9 and UGT2B7 but not UGT1A1 17; however, accumulation of mycophenolate is explained by the mutation involving exon 4 of the UGT1A1 gene and thus affecting all UGT1A isoforms 18. Second, 6 mo after transplantation, patient 1 presented with crusted scabies ( Scabies norvegica ) infection, a particularly severe form found in immune‐compromised patients.…”

Analysis of the UDP-glucuronosyltransferase gene in Portuguese patients with a clinical diagnosis of Gilbert and Crigler–Najjar syndromes

“…For this reason, our patient could have a more severe phenotype than the patients homozygous for the [TA]7 allele. Erps et al [36] first described the c.923G>A mutation in a 7-year-old female with Crigler -Najjar syndrome type I. This mutation, coincident with a highly conserved residue in the protein sequence, results in the lack of enzyme activity.…”