1. Sponsor Organization: NIH/NIEHS

2. Project Title: 07685-03 RECOMBINANT DETECTION SYSTEMS FOR DIOXIN LIKE CHEMICALS

3. Project Focus:

• Project Primary Focus: Exposure Assessment
• Project Secondary Focus:

4. Description:

Halogenated aromatic hydrocarbons (HAHs), such as polychlorinated-dibenzo-p-dioxins (PCDDs), PCBs, and dibenzofurans (PCDFs) produce a wide variety of toxic effects. Although exposure to specific HAHs, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD,dioxin), can produce these effects at low concentrations, the induction of cytochrome P45O1A1 by HAHs is one response that is highly conserved across species and is used as the model system for analysis of the mechanism of action of HAHs. Induction of P45O1A1 is mediated by a soluble intracellular protein (the Ah receptor (AhR)) which binds the HAH specifically and with high affinity. After binding, HAH:AhR complexes are converted into their DNA binding form, they subsequently accumulate within the nucleus and activate gene transcription through a high affinity interaction with a specific DNA sequence (dioxin responsive enhancers) upstream of the P45O1A1 gene. In addition to mediating the induction of P45O1A1, structure-activity relationship studies have demonstrated a strong correlation between the ability of a chemical to bind to the AhR and its ability to produce toxicity, implicating the AhR in mediating the toxicity of HAHs. Thus, many, if not all, of the toxic and biological responses to HAHs currently appear to mediated by the AhR. This project will use several mechanistic aspects of this AhR-dependent system to develop bioassay systems for the detection of HAHs. Given the high degree of correlation between the ability of a chemical to bind to the AhR, its ability to induce AhR transformation and DNA binding and gene expression, we will develop two AhR DNA binding assays (gel retardation analysis and a novel membrane filtration assay) as bioassays for detection of dioxin-like chemicals. In addition, we will stably transfect several HAH-inducible expression vectors which contains a luciferase or alkaline phosphatase reporter gene, into HAH-responsive human and mouse cell lines. Exposure of these recombinant cell lines to dioxin-like HAHs will result in the induction of expression of the reporter gene to a level proportional to the HAH dose.

5. References:

6. Inventory Category:

• Primary: Methods
• Secondary:

7. Inventory Subcategory:

• Primary: Exposure and Risk Models
• Secondary:

8. Keywords for Experimental System/Species:

• Species:
  • Mammal
  • Human
  • Mouse
• Study Type:
  • In Vitro
  • Laboratory Study
• Fate and Transport:

9. Keywords for Experimental Endpoints:

• Health Effect:
• Hormonal Measures:
  • Ah Receptor
• Level Of Study:
  • Gene Expression
• Chemistry Metabolism:
  • Cytochrome P450
• Life Stage:
• Risk Assessment:

10. Chemical Agents:

• Dioxins
• Pcbs
• Pcdfs

11. Performing Institution:

• UNIVERSITY OF CALIFORNIA DAVIS

12. Contact:

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