Prenatal diagnosis of bilirubin-UDP-glucuronosyltransferase deficiency in rats by genomic DNA analysis.
Hepatic bilirubin excretion requires UDP-glucuronosyltransferase-mediated glucuronidation. Patients with type I Crigler-Najjar syndrome and mutant rats (Gunn strain) inherit deficiency of UDP-glucuronyltransferase activity toward bilirubin as an autosomal recessive trait and, as a result, exhibit marked nonhemolytic unconjugated hyperbilirubinemia throughout postnatal life. Heterozygous carriers of the trait have normal serum bilirubin levels. Because of placental excretion of unconjugated bilirubin, type 1 Crigler-Najjar syndrome patients and Gunn rats are not jaundiced in utero, making prenatal diagnosis difficult. Here we report a diagnostic method in Gunn rats based on genomic DNA analysis for prenatal recognition of deficiency of UDP-glucuronyltransferase activity toward bilirubin in Gunn rats and identification of heterozygous carriers. We and others have shown that two distinct messenger RNA species (UDP-glucuronyltransferase activity toward bilirubin and the 3-methylcholanthrene-inducible phenol-UDP-glucuronyltransferase messenger RNA) in Gunn rat liver contain identical deletions of a single guanosine residue in their common 3' regions. Loss of the restriction site for the endonuclease BstNI, which results from this deletion, was used as the basis for a diagnostic test. Female heterozygous Gunn rats were mated with male homozygous Gunn rats. Genomic DNA was extracted from the chorionic aspect of placenta of 17-day fetuses or from leukocytes from normal rats, obligate heterozygotes and homozygous Gunn rats. The DNA was sequentially digested with the restriction enzymes EcoRI and BstNI and subjected to Southern-blot analysis with a double-stranded DNA probe for the common region of UDP-glucuronyltransferase activity toward bilirubin and the 3-methylcholanthrene-inducible UDP-glucuronyltransferase messenger RNAs.(ABSTRACT TRUNCATED AT 250 WORDS)
Huang
T J
TJ
Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, New York 10461.
Chowdhury
J R
JR
Lahiri
P
P
Yerneni
P C
PC
Bommineni
V R
VR
Arias
I M
IM
Chowdhury
N R
NR
D000818:Animals; D001483:Base Sequence; D004247:DNA / Q000032:analysis*; D006580:Genetic Carrier Screening; D005838:Genotype; D005964:Glucosyltransferases / Q000172:deficiency* / Q000235:genetics; D006579:Heterozygote; D006720:Homozygote; D008969:Molecular Sequence Data; D010641:Phenotype; D016133:Polymerase Chain Reaction; D011296:Prenatal Diagnosis / Q000379:methods*; D012333:RNA, Messenger / Q000032:analysis*; D051381:Rats; D011911:Rats, Gunn; D011919:Rats, Inbred Strains
D012333:RNA, Messenger; D004247:DNA; D005964:Glucosyltransferases; C035814:bilirubin UDP-glucosyltransferase