An Aryl Hydrocarbon Receptor Sequence from the Caecilian Gymnopis multiplicata Exhibits Low Responsiveness to 2,3,7,8-tetrachlorodibenzo-p-dioxin

The aryl hydrocarbon receptor (AHR) mediates the toxic effects of dioxin-like compounds, such as 2,3,7,8-tetracholordibezno-p-dioxin (TCDD). Sequence and structural features of AHR can underlie species- and population-specific differences in its affinity for TCDD and other agonists. While most vertebrates possess at least one AHR that binds TCDD tightly, all characterized amphibian AHRs bind TCDD with very low affinity. Our previous analyses of AHRs from Xenopus laevis (a frog; order Anura) and Ambystoma mexicanum (a salamander; order Urodela) identified three key amino acid residues in the ligand binding domain (LBD) that underlie low affinity binding. In X. laevis AHR1β, these are A354, A370, and N325. In studying the AHR of the caecilian Gymnopis multiplicata, we sought to determine if AHRs from all three amphibian orders share the low affinity phenotype. G. multiplicata represents Order Apoda (caecilians), a clade of legless amphibians that diverged early from the class’s common lineage, prior to the split between frogs and salamanders. cDNA was isolated by RT-PCR from tissue provided by the Museum of Vertebrate Zoology at the University of California, Berkeley. The encoded protein (92.7 kDa) is monophyletic with vertebrate AHR1s, sharing 59% identity with X. laevis AHR1β and 63% identity with A. mexicanum AHR. An LBD homology model suggests that the basic architecture of the caecilian AHR closely resembles other species, and the LBD sequence includes the three signature residues of low TCDD affinity. We measured the TCDD responsiveness with transactivation assays employing pGudLuc6.1, a luciferase reporter construct driven by the mouse CYP1A1 enhancer region. For G. multiplicata AHR, the EC50 for reporter gene induction by TCDD was 28.46 nM, similar to X. laevis AHR1β (26.84 nM) and dramatically less than a chimeric frog AHR containing the mouse LBD (0.2 nM). Taken together, low TCDD responsiveness and sequence conservation with the frog and salamander AHRs suggest that this caecilian AHR also binds TCDD with low affinity. We predict G. multiplicata is similarly resistant to the toxic effects of TCDD and other xenobiotic AHR agonists. (NIH: R15ES011130, P42ES007381, R01ES007685)