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Event Title

ARNT/HIF1-alpha, reduced dimerization

Key Event Overview

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AOPs Including This Key Event

AOP Name Event Type Essentiality
Aryl hydrocarbon receptor activation leading to embryolethality via cardiotoxicty KE Moderate

Taxonomic Applicability

Name Scientific Name Evidence Links

Level of Biological Organization

Biological Organization

How this Key Event works

Sustained dimerization of the aryl hydrocarbon receptor (AHR) and AHR nuclear translocator (ARNT) induced by xenoiotics may sequester ARNT from its other dimerization partners at inappropriate times during embryonic cardiomorphogenesis, disrupting ARNT-dependent cellular functions (Heid et al. 2001; Walker et al. 1997). ARNT serves as a dimerization partner for hypoxia inducible factor 1&alph; (HIF-1α), and this complex is involved in mediating physiological responses to hypoxia. Dimerization between ARNT and HIF-1α forms a transcription factor complex (HIF-1) that binds to hypoxia response enhancer sequences on DNA to activate the expression of genes such as vascular endothelial growth factor (VEGF), which is involved in angiogenesis (Forsythe et al. 1996; Goldberg and Schneider 1994; Jiang et al. 1996; Maxwell et al. 1997; Shweiki et al. 1992).

How it is Measured or Detected

Methods that have been previously reviewed and approved by a recognized authority should be included in the Overview section above. All other methods, including those well established in the published literature, should be described here. Consider the following criteria when describing each method: 1. Is the assay fit for purpose? 2. Is the assay directly or indirectly (i.e. a surrogate) related to a key event relevant to the final adverse effect in question? 3. Is the assay repeatable? 4. Is the assay reproducible?

The active HIF1- α complexed with ARNT can be measured using protein-DNA interaction assays. Two methods are described in detail by Perez-Romero and Imperiale (Perez-Romero and Imperiale 2007). Chromatin immunoprecipitation measures the interaction of proteins with specific genomic regions in vivo. It involves the treatment of cells with formaldehyde to crosslink neighboring protein-protein and protein-DNA molecules. Nuclear fractions are isolated, the genomic DNA is sheared, and nuclear lysates are used in immunoprecipitations with an antibody against the protein of interest. After reversal of the crosslinking, the associated DNA fragments are sequenced. Enrichment of specific DNA sequences represents regions on the genome that the protein of interest is associated with in vivo. Electrophoretic mobility shift assay (EMSA) provides a rapid method to study DNA-binding protein interactions in vitro. This relies on the fact that complexes of protein and DNA migrate through a non-denaturing polyacrylamide gel more slowly than free DNA fragments.

Evidence Supporting Taxonomic Applicability


1. Forsythe, J. A., Jiang, B. H., Iyer, N. V., Agani, F., Leung, S. W., Koos, R. D., and Semenza, G. L. (1996). Activation of vascular endothelial growth factor gene transcription by hypoxia-inducible factor 1. Mol. Cell Biol. 16(9), 4604-4613.

2. Goldberg, M. A., and Schneider, T. J. (1994). Similarities between the oxygen-sensing mechanisms regulating the expression of vascular endothelial growth factor and erythropoietin. J. Biol. Chem. 269(6), 4355-4359.

3. Heid, S. E., Walker, M. K., and Swanson, H. I. (2001). Correlation of cardiotoxicity mediated by halogenated aromatic hydrocarbons to aryl hydrocarbon receptor activation. Toxicol. Sci 61(1), 187-196.

4. Jiang, B. H., Rue, E., Wang, G. L., Roe, R., and Semenza, G. L. (1996). Dimerization, DNA binding, and transactivation properties of hypoxia-inducible factor 1. J. Biol. Chem. 271(30), 17771-17778.

5. Maxwell, P. H., Dachs, G. U., Gleadle, J. M., Nicholls, L. G., Harris, A. L., Stratford, I. J., Hankinson, O., Pugh, C. W., and Ratcliffe, P. J. (1997). Hypoxia-inducible factor-1 modulates gene expression in solid tumors and influences both angiogenesis and tumor growth. Proc. Natl. Acad. Sci U. S. A 94(15), 8104-8109.

6. Shweiki, D., Itin, A., Soffer, D., and Keshet, E. (1992). Vascular endothelial growth factor induced by hypoxia may mediate hypoxia-initiated angiogenesis. Nature 359(6398), 843-845.

7. Walker, M. K., Pollenz, R. S., and Smith, S. M. (1997). Expression of the aryl hydrocarbon receptor (AhR) and AhR nuclear translocator during chick cardiogenesis is consistent with 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced heart defects. Toxicol. Appl. Pharmacol. 143(2), 407-419.