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EDRI Federal Project Inventory:
61262-02 Metallothionein in Reproduction and Development
- Sponsor Organization: NIH/NCI
- Project Title: 61262-02 METALLOTHIONEIN IN REPRODUCTION AND DEVELOPMENT
- Project Focus: HUMAN HEALTH EFFECTS
- Description: These studies address research objectives of this RFA including:
mechanisms of regulation of metallothionein (MT)gene expression, metal
induction during development, and organs specificity in transgenic
models. The overallobjective of theses studies is to understand the
regulation and functions of MT in reproduction and
embryonicdevelopment in mammals. This information, in turn, is likely
to provide important clues as to possible roles of MT
incarcinogenesis. Mt plays a pivotal role in essential metal
homeostasis and in protection from toxic metals. Zinc (Zn),copper
(Cu), and cadmium (Cd) have dramatic effects on embryogenesis.
Insufficient dietary Zn or Cu leads toabnormal development and
reproductive failure, whereas Cd, is embryotoxic, teratogenic, and
carcinogenic. Our studieson mouse embryogenesis establish that the MT
genes are actively expressed in specific cell-types that surround
thedeveloping embryo from the time of implantation to near
parturition, as well as in metal-treated reimplantationblastocysts and
in fetal hepatocyte at midgestation. This suggests that MT has
important functions duringembryogenesis. The specific aims of the
proposed studies are to: 1) determine mechanisms regulating the cell-
typespecific expression of these genes in the reproductive tract and
embryo, and 2) delineate the functions of these proteinsduring
embryogenesis. Specific aim 1 will be approached by; a) examining the
effects of well-documented MT genetranscriptional regulators on MT
gene expression in vivo and/or in vitro in mouse decidual cells,
visceral endoderm,reimplantation embryos and fetal hepatocytes, b)
determining the in vivo interactions of proteins with the MT-I
genepromoter in specific cell-types, using genomic footprinting
techniques, and c) ultimately mapping cis-acting promotersequences
involved in MT gene expression in these cell-types using transgenic
mouse strains that express reporter genesunder the control of MT-I
promoter deletion mutants. A testable hypothesis is that tissue-
specific enhancer elementsexist in the mouse MT-I promoter.
Delineation of MTs functional roles in vivo is hampered by a lack of
strains of micethat exhibit defects (mutations) effecting MT gene
expression. Thus, specific aim 2 will be approached by
creatingstrains of transgenic mice that; a) display constitutively
heightened, or b) misexpression of the MT genes. Transgenicmice that
constitutively over-express MT, due to integration multiple copies of
the intact mouse MT-I gene, are currentlyunder study. Transgenic
strains that over-express the MT gene in the pancreas, liver and
uterus due to enhancer-driventransgene expression will be created.
Gene expression will be determined at the level of mRNA by Norther, in
situ andsolution hybridization, and the reverse transcriptase-
polymerase chain reaction. MT protein synthesis and accumulationwill
be determined by the Cd-heme assay, pulse-labeling western blotting,
immunocytochemistry, and anion exchangechromatography. Transgenic
mice will be analyzed for effects of aberrant MT gene expression on
normalembryogenesis as well as on the embryos response to exogenous
metals. A testable hypothesis is that misexpression ofMT will alter
metal homeostasis during pregnancy causing increased sensitivity to
dietary Zn deficiency and reducedsensitivity to Cd toxicity.
- References:
- Category: MODELS
- Subcategory: BASIC RESEARCH
- Keywords for Experimental System/Species: IN VITRO, IN VIVO, MAMMALIAN
- Keywords for Experimental Endpoints: CARCINOGENESIS, EMBRYO, GENE EXPRESSION, TRANSGENIC MODEL
- Chemical Agents: CADMIUM, LEAD
- Performing Institution: UNIVERSITY OF KANSAS MEDICAL CENTER
- Contact: CONTRACT PERSON: ELAINE C. LEE; BUILDING 31; 11A21; NATIONAL CANCER
INSTITUTE, NIH,BETHESDA, MD 20892-2590; 301-496-5515;
LEEE@OD.NCI.NIH.GOV
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