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Molecular Biology, Environmental Toxicology

Our research seeks to understand the molecular mechanisms of contaminant toxicity in aquatic and marine animals.

Recent publications (underline indicates Kenyon student co-author):

Laub, Leo B. Brian D. Jones, and Wade H. Powell (2010) Responsiveness of a Xenopus laevis cell line to the aryl hydrocarbon receptor ligands 6-formylindolo[3,2-b]carbazole (FICZ) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) Chem.Biol.Interact., 83(1):202-11

Anna L. Zimmermann, Elizabeth A. King, Emelyne Dengler, Shana R. Scogin, and Wade H. Powell (2008) An Aryl Hydrocarbon Receptor Repressor from Xenopus laevis: Function, Expression and Role in Dioxin Responsiveness during Frog Development. Toxicol. Sci. 104:124-134.

Philips, B.H., T.C. Susman, and W.H. Powell. (2006) Differences in elimination of 2,3,7,8-tetrachlorodibenzo-p-dioxin during Xenopus laevis development. Marine Environmental Research, 62:S34-S37.

Lavine, J.A., A.J. Rowatt, T. Klimova, A.J. Whitington, E. Dengler, C. Beck, and W.H. Powell (2005) Aryl Hydrocarbon Receptors in the frog Xenopus laevis: Two AHR1 paralogs exhibit low affinity for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) Toxicol. Sci. 88:62-70.

Rowatt, A.J., J.J. DePowell, and W.H. Powell. (2003) ARNT gene multiplicity in amphibians: Characterization of ARNT2 from the frog, Xenopus laevis. Journal of Experimental Zoology Part B: Molecular and Developmental Evolution, 300B:48 - 57

 

Kenyon Lab Group Summer 2010
Wade Powell, Cami Odio '11, Chad Kurylo '11


Multiple low-affinity aryl hydrocarbon receptors from the frog Xenopus laevis

Embryos of the African clawed frog, Xenopuslaevis, represent a long standing model of vertebrate development.  They are also used in FETAX (Frog Embryos Teratogenesis Assay-Xenopus) and similar assays of the developmental toxicity of chemicals and environmental samples.  2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD) is a potent developmental toxicant in most vertebrates.  However, frogs are generally insensitive to TCDD toxicity, especially at early life stages.  Thus, FETAX and other frog embryo toxicity tests may be poorly suited for determining the developmental toxicity of samples containing dioxin-like compounds.  Our research seeks to delineate the molecular mechanisms underlying TCDD insensitvity in frogs, using X. laevis as a model system. 

The aryl hydrocarbon receptor (AHR) is a ligand activated transcription factor that mediates the biological and toxicological effects of a broad range of structurally diverse chemicals, including TCDD. Inherent properties of the AHR signaling pathway, including AHR expression levels and the affinity of AHR for specific ligands, can underlie large variations in the relative potency of different ligands and the sensitivity of different animal groups. However, neither the exact mechanisms by which AHR activity leads to toxicity nor the endogenous, non-toxicological functions of AHR signaling are well understood. 

Our group has identified two AHRs from the African clawed frog (Xenopus laevis), recently duplicated paralogs called AHR1a and AHR1b.  Both proteins bind TCDD with at least 25-fold lower affinity than the AHR from a highly sensitive strain of mouse, likely accounting for the dioxin-insensitive phenotype. Our research takes advantage of the unique amino acid sequence, functional properties, and phylogenetic position of the frog AHRs to probe their structural interactions with a range of xenobiotic and naturally occurring ligands and to contrast their function with AHRs from TCDD-sensitive species and with each other.  We have three main goals:

  1. Using site-directed mutagenesis to make the frog AHRs more “mouse-like,” we will test the hypothesis that changes in one or a few amino acids within the putative ligand binding domain confer low TCDD affinity. This comparative approach will contribute significantly to the identification of important structural features of AHR’s ligand binding pocket.
  2. We will determine the relative potency of structurally diverse candidate ligands.  Although X.laevis AHRs bind TCDD with low affinity, they may remain highly responsive to structurally distinct compounds, especially putative endogenous ligands. We will test this hypothesis by establishing structure-activity relationships for a range of candidate ligands, including indole-containing compounds that bind mammalian AHRs. 
  3. We will determine the functional differences between AHR1a and AHR1b, examining expression patterns, enhancer preferences, and broad-based changes in gene expression mediated by individual paralogs.  These studies will test the hypothesis that the AHR paralogs exhibit distinct functions, possibly partitioning multiple roles of the single mammalian AHR.

Overall, this comparative approach in a novel model system will provide important basic information about the structure and function of all vertebrate AHRs. Understanding the differences between frog and human AHRs will also aid risk assessment by refining interpretation of toxicological data derived from FETAX (Frog Embryo Teratogenesis Assay-Xenopus) and similar developmental toxicity tests that employ frog embryos. development.

[Supported by the National Institutes of Health, R15-ES011130]


2,3,7,8 tetrachlorodibenzo-p-dioxin
("TCDD" or "dioxin")


Xenopus laevis
adult female


Xenopus laevis
embryos
(frog photos from www.xlaevis.com)