AHR is a cytoplasmic receptor that has affinity for numerous xenobiotic ligands, which include halogenated aromatic hydrocarbons such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), one of the most potent AHR activators, and AHR signaling mediates the detrimental effects of environmental contaminants on body tissues and organs. Ligand binding induces nuclear translocation of AHR and interaction with the AHR binding partner, aryl hydrocarbon receptor nuclear translocator (ARNT), allowing recognition of specific DNA enhancer sequences. More recently endogenous and natural AHR ligands, such as tryptophan metabolites and dietary compounds, have been identified. These ligands induce AHR-mediated immunomodulatory effects such as controlling normal T cell differentiation and immune tolerance. However, little is known about the role of ARNT in AHR immune signaling. Initial studies by our laboratory, and others, into the regulatory role of ARNT in AHR-mediated immunomodulation have suggested that ARNT is an integral cofactor in different immune signaling pathways, including AHR and NF-?B signaling. ARNT is often described in AHR signaling as a constitutively expressed, non-regulated, nuclear binding partner for AHR. However, our data challenge this assumption and show that the actual regulatory paradigm is more intricate. For instance, ARNT is expressed as two isoforms, isoform 1 and 3, which differ in only 15 amino acids present in isoform 1. Despite their sequence similarity, we have found that the ARNT isoforms appear to have opposing functions in AHR signaling. Intriguingly, the extra 15 amino acids in isoform 1 include a unique phosphorylation site, where we have observed TCDD-induced phosphorylation. Furthermore, we find that the phosphorylation of ARNT isoform 1 is crucial for the recruitment of RNA polymerase II and transactivation of AHR/ARNT target genes. We predict that ARNT activity is a function of both unique phosphorylation of isoform 1 and the given isoform ratio within a particular cell type, which in turn is likely important for regulating the magnitude/outcome of the AHR response. To investigate our hypothesis we propose to 1) Define a comprehensive molecular framework for AHR regulation by the ARNT isoforms, 2) Examine the role of AHR activity, as governed by the ARNT isoforms, in lymphoma cell growth, and 3) Characterize the regulation of AHR signaling by ARNT isoforms in vivo. Ultimately, these studies will lay the groundwork for ARNT-based therapies such as splice modulation as a means of manipulating the ARNT isoforms to control AHR signaling in autoimmune diseases and cancer.

Public Health Relevance

The experiments outlined in this proposal are focused on deciphering the regulatory roles of ARNT isoforms on AHR signaling in immune cell biology. While ARNT is a known AHR binding partner, ARNT activity in the cell has not been considered as the sum of two isoforms, which we find differentially regulate AHR function. Given the importance of AHR to hematopoiesis, and lymphocyte differentiation and function, it is expected that this work will provide a rational basis for the future development of ARNT and AHR-based therapies for immune modulation in autoimmune disease and cancer.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Project (R01)
Project #
7R01ES025809-02
Application #
9477928
Study Section
Systemic Injury by Environmental Exposure (SIEE)
Program Officer
Humble, Michael C
Project Start
2016-06-01
Project End
2021-05-31
Budget Start
2017-06-01
Budget End
2018-05-31
Support Year
2
Fiscal Year
2017
Total Cost
Indirect Cost
Name
University of Texas Med Br Galveston
Department
Pharmacology
Type
Schools of Medicine
DUNS #
800771149
City
Galveston
State
TX
Country
United States
Zip Code
77555