Project Details

Description

UDP-glucuronosyltransferases (UGT) are phase II xenobiotic metabolizing
enzymes that glucuronidate a variety of exogenous and endogenous
compounds. This family of enzymes play an important role in the
metabolism of many carcinogens such as hydroxylated benzo[a]pyrene,
ben[a]anthracene, 2,3,7,8-tetrachlorobenzo-p-dioxin (TCDD), aflatoxin B1,
acetylaminofluorene, and tobacco-specific nitrosamine such as 4-
(methylnitrosamino)-1-(3-pyridyl)-butan-1-ol NNAL). This proposal
focuses on the isolation and characterization of the genes encoding for
two major types of UDP-glucuronosyltransferases, i.e., bilirubin/phenol
(UGT-br/p) and bile acid/steroid (UGT-ba/s) UGT in mice. The underlying
hypothesis is that cloning, gene expression and regulation studies will
offer unique insights into the biological mechanisms of detoxification
and/or toxicity of different carcinogens catalyzed by UGTs in the mouse
and will aid in the extrapolation of chemical toxicity, carcinogenicity
and mutagenicity studies from mouse to human. The specific aims of this
research proposal are: 1) To clone, sequence and characterize the cDNAs
encoding for the mouse bilirubin/phenol (UGTbr/p) and the bile
acid/steroid (UGTba/s) UDP-glucuronosyltransferases. This will involve
a series of experiments aimed at screening and re-screening of a female
mouse liver cDNA library using human and mouse cDNA probes, followed by
restriction-mapping and sequencing. 2) To elucidate the catalytic
activities and toxicology of mouse UGT via cDNA expression in cultured
cells. This will involve the expression of the cloned mouse UGTbr/p and
UGTba/s cDNAs in fibroblast cell lines and baculovirus system and
assaying for glucuronidation activities towards model compounds as well
as known carcinogenic substrates. 3) To characterize the
regulation/expression of UGTs in hepatoma cell lines and in whole
animals. These experiments will involve administration of known inducers
for Phase II drug metabolizing enzymes as well as environmental
pollutants such as TCDD and DDT (1,1-bis[p-chlorophenyl]-2,2,2-
trichloroethane) to: 2) hepatoma cell lines - Hepa 1c1c7 (mouse), Fao
(rat) and Hep G2 (human); and b) different strains and gender of mice -
Balb/c, C3H, C57BL/6J, B6C3F1, ICR and DBA/2J mice, and assaying for mRNA
levels as well as UGT activities toward model and carcinogenic
substrates. 4) To isolate, sequence and characterize the genomic DNA of
the mouse UGTbr/p and UGTba/s genes to elucidate the conserved domain(s)
at the 5' flanking regions of the genes. This involves screening of a
mouse genomic library with their respective cloned mouse UGTbr/p and
UGTba/s cDNAs and subsequent restriction-mapping and sequencing two kb
5' flanking regions of the genes. The results of the proposed
experiments are likely to significantly enhance our fundamental
understanding of the biological mechanisms of UGTs in the glucuronidation
and detoxification of carcinogens in mice and may lend insights into
human carcinogenesis and toxicity induced by chemical and environmental
carcinogens.Description
StatusFinished
Effective start/end date7/1/946/30/98

Funding

  • National Institutes of Health: $113,802.00
  • National Institutes of Health
  • National Institutes of Health: $18,000.00
  • National Institutes of Health

ASJC

  • Environmental Science(all)
  • Medicine(all)

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