The role of DNA-binding and ARNT dimerization on the nucleo-cytoplasmic translocation of the aryl hydrocarbon receptor

Rashad Haidar,Aline Rosin,Peter Laux,Doris Genkinger,Josephine Kugler,Andreas Luch,Frank Henkler

doi:10.1038/s41598-021-97507-w

Rashad Haidar, Aline Rosin + Show 5 more

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https://doi.org/10.1038/s41598-021-97507-w

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Abstract

The human aryl hydrocarbon receptor (AHR) is predominantly located in the cytoplasm, while activation depends on its nuclear translocation. Binding to endogenous or xenobiotic ligands terminates the basal nucleo-cytoplasmic shuttling and stabilizes an exclusive nuclear population. The precise mechanisms that facilitate such stable nuclear accumulation remain to be clarified as essential step in the activation cascade. In this study, we have tested whether the sustained nuclear compartmentalization of ligand-bound or basal AHR might further require heterodimerization with the AHR-nuclear translocator (ARNT) and binding to the cognate XRE-motif. Mutagenesis of the DNA-binding motif or of selected individual residues in the ARNT-binding motif did not lead to any variation in AHR’s nucleo-cytoplasmic distribution. In response to ligands, all mutants were retained in the nucleus demonstrating that the stable compartmentalization of activated AHR in the nucleus is neither dependent on interactions with DNA, nor ARNT. Knocking down the ARNT gene using small interfering RNA confirmed that ARNT does not play any role in the intracellular trafficking of AHR.

Highlights

The human aryl hydrocarbon receptor (AHR) is predominantly located in the cytoplasm, while activation depends on its nuclear translocation
We have investigated whether the ligand-induced nuclear accumulation of the AHR depends on its heterodimerization with AHR-nuclear translocator (ARNT) and its DNA-binding domain that associates with the cognate xenobiotic response elements (XRE) motif
To design AHR mutants that are incapable of interacting with ARNT

Summary

Introduction

The human aryl hydrocarbon receptor (AHR) is predominantly located in the cytoplasm, while activation depends on its nuclear translocation. All mutants were retained in the nucleus demonstrating that the stable compartmentalization of activated AHR in the nucleus is neither dependent on interactions with DNA, nor ARNT. The aryl hydrocarbon receptor (AHR) is a ligand induced transcription factor that activates oxidative phase I metabolism by the induction of cytochrome P450-dependent monooxygenases (CYP), especially CYP1A1 and 1B11. Endogenous AHR agonists include tryptophan metabolites, such as kynurenine (KYN), and indole derivatives, such as indirubin (IND)[14] The latter is confirmed to be present in the urine of healthy people with levels sufficient to activate the AHR15. Transcriptional activation of the AHR is a tightly regulated process that depends on the interactions with several co-factors and the heterodimerization with the AHR-nuclear translocator (ARNT). The DNA-binding motif of the AHR overlaps with the NLS, more accurately with the amino acids Histidine (H39) and Arginine (R40) in the second N LS22–24

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