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Cited by 15Toxic Organic Pollutants ImpactPharmacogenetics and Drug MetabolismCarcinogens and Genotoxicity Assessment

Generation of an Allelic Series at the Ahr Locus Using an Edited Recombinant Approach

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Wilson, Rachel H, Carney, Patrick R, Glover, Edward, Parrott, Jessica C, Rojas, Brenda L, Moran, Susan M, Yee, Jeremiah S, Nukaya, Manabu +5 more·2021·Toxicological Sciences
DOI 10.1093/toxsci/kfab005PMC PMC8041461
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Abstract

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor and a member of the PER-ARNT-SIM (PAS) superfamily of environmental sensors. The AHR is involved in a series of biological processes including adaptive metabolism of xenobiotics, toxicity of certain environmental pollutants, vascular development, fertility, and immune function. Mouse models, including the Ahr null and Ahr conditional null (Ahrfx) mice, are widely used for the study of AHR-mediated biology and toxicity. The Ahr conditional null mouse harbors the low-affinity Ahrd allele that exhibits approximately a 10-fold lower binding affinity for certain xenobiotic AHR ligands than the widely used C57BL/6 mouse that harbors the higher affinity Ahrb1 allele. Here, we report a novel mouse model that introduces a V375A polymorphism that converts the low-affinity allele into a high-affinity allele, offering a more sensitive conditional model. In the generation of this novel conditional allele, two additional mutants arose, including a 3-bp deletion in the PAS-B domain (AhrNG367R) and an early termination codon in the PAS-B domain (AhrTer383). The AhrNG367R allele presents as a phenocopy of the null and the AhrTer383 allele presents as an antimorph when assessing for the ductus venosus and liver lobe weight endpoints. These new models represent a series of tools that will be useful in further characterizing AHR biology.

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  • Excision of AhrTer383 in the endothelial compartment of AhrTer383CreCdh5 mice rescues the patent DV phenotype, with 16% penetrance.

    “Yet, excision of the AhrTer383 allele in the endothelial compartment (CreCdh5 ) of AhrTer383CreCdh5 mice rescues the patent DV phenotype (16%) (R. H. Wilson et al., 2021).”

    From Rodent Genetic Models of Ah Receptor Signaling (2021) · cites this paper as R. H. Wilson et al., 2021

    The Results (Table 5) show that endothelial excision of AhrTer383 reduced patent DV in heterozygous mice from 86% (19/22) to 18% (2/11), supporting the rescue claim. The cited 16% penetrance does not match the paper’s reported 18%. (LLM, conf=0.92)

    Experimental — accuracy not yet measured

  • AHRTer383 is not expected to bind ligand because the frameshift is near the LBD, but its NLS, DBD, and ARNT-binding domains remain intact and are expressed at 37 kDa.

    “Due to the proximity of the frame-shift mutations to the LBD, AHRTer383 is not expected to bind ligand, though the NLS, DBD, and ARNT binding domains remain intact and are expressed at 37 kDa, as demonstrated in protein expression experiments (Dolwick, Swanson, et al., 1993; R. H. Wilson et al., 2021; Xing et al., 2012)).”

    From Rodent Genetic Models of Ah Receptor Signaling (2021) · cites this paper as R. H. Wilson et al., 2021

    The Discussion (page not provided) describes the frameshift, disrupted ligand-binding pocket, retained bHLH and PAS-A domains, and predicted ARNT dimerization and DNA interaction. It also reports an approximately 37 kDa protein, consistent with the Results protein-expression section. The copy does not establish that AHRTer383’s NLS remains intact; ARNT and DNA interaction are predictions, not demonstrated by the protein-expression experiment. (LLM, conf=0.83)

    Experimental — accuracy not yet measured

  • Preliminary modeling suggests the PAS B domain in AhrNG367R may be misfolded because glycine G368, critical for proper folding, is removed.

    “Preliminary modeling studies suggest that the PAS B domain may be misfolded due to the removal of a glycine residue (G368) that is critical to the proper folding of this region (R. H. Wilson et al., 2021).”

    From Rodent Genetic Models of Ah Receptor Signaling (2021) · cites this paper as R. H. Wilson et al., 2021

  • Mice homozygous for AhrNG367R are a functional phenocopy of AHR null mice in hepatovascular development and gene induction.

    “This prediction is supported by the observation that mice homozygous for this allele are a functional phenocopy of the null in hepatovascular development and gene induction studies (R. H. Wilson et al., 2021).”

    From Rodent Genetic Models of Ah Receptor Signaling (2021) · cites this paper as R. H. Wilson et al., 2021

  • AhrTer383 arose as an apparent CRISPR-mediated NHEJ artifact during attempts to generate AhrV375A.

    “Like the AhrNG367R allele described above, the AhrTer383 mutant appears to have been generated as an artifact of CRISPR-mediated NHEJ during the effort to generate AhrV375A (Figure 5)(Ceccaldi et al., 2016; R. H. Wilson et al., 2021).”

    From Rodent Genetic Models of Ah Receptor Signaling (2021) · cites this paper as R. H. Wilson et al., 2021

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Authors
Wilson, R. H., Carney, P. R., Glover, E., Parrott, J. C., Rojas, B. L., Moran, S. M., Yee, J. S., Nukaya, M., Goetz, N. A., Rubinstein, C. D., Krentz, K. J., Xing, Y., & Bradfield, C. A.
Affiliations
  • University of Wisconsin–Madison (US)
  • Biotechnology Center, University of Wisconsin–Madison, Madison, Wisconsin 53706, USA
Corresponding author
Bradfield, Christopher A
ORCID
Venue
Toxicological Sciences
Publisher
Oxford University Press (OUP)
Published
2021-01-22
Volume
180
Issue
2
Pages
239-251
ISSN
  • 1096-0929 (ISSN-L)
  • 1096-6080
DOI
10.1093/toxsci/kfab005
PMC
PMC8041461
OpenAlex
W3122690459
Cited by
15
References
11
Citations by year
2026: 1 · 2025: 5 · 2024: 3 · 2022: 2 · 2021: 4
FWCI
0.89
Type
journal article
Language
en
Open access
Licence
academic.oup.com/journals/pages/open_access/funder_policies/chorus/standard_publication_model (vor)
Locations
  • Toxicological Sciences (publishedVersion)
  • PubMed Central (publishedVersion)
  • PubMed (publishedVersion)
Funders
  • National Institutes of Health (R35-ES028377, T32-ES007015, T32-GM008692, T32-CA009135, P30-CA014520)
  • The UW SciMed GRS Program
  • The Morgridge Foundation
Topics
  • Physical Sciences › Environmental Science › Health, Toxicology and Mutagenesis › Toxic Organic Pollutants Impact
  • Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Pharmacology › Pharmacogenetics and Drug Metabolism
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Carcinogens and Genotoxicity Assessment
Keywords
aryl hydrocarbon receptor, allelic series, PAS-B domain, phenocopy, ductus venosus, xenobiotic metabolism, environmental pollutants, transcription factors
MeSH terms
Alleles, Animals, Liver, Mice, Inbred C57BL, Receptors, Aryl Hydrocarbon, Mice, Knockout, Mice, Aryl Hydrocarbon Receptor Nuclear Translocator, Basic Helix-Loop-Helix Transcription Factors, Basic Helix-Loop-Helix Proteins

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