AbstractThe Aryl Hydrocarbon Receptor (AhR) and the Aryl Hydrocarbon Nuclear Translocator (ARNT) play a role in mediating transcriptional responses to environmental pollutants, including the highly toxic compound 2,3,7,8-tetrachlorodibenzo -p-dioxin (TCDD) but also endogenous physiological ligands. More recent studies have also indicated that the AhR plays a role in the immune system notably in effector Th17 cells where it seems to be critical for the production of the IL-22 cytokine. It is known that AhR ligands such as dioxins can suppress CD8 T cell mediated antiviral immune responses but it is not known whether this reflects a direct role of the AhR in CD8 T cells.
Accordingly, one objective of the present study was to explore AhR and ARNT expression in CD8 T cells. The initial strategy was to probe AhR and ARNT expression by western blot analysis. A second approach was to develop a mouse model that would fate mark single lymphocytes that have activated AhR signaling pathways. A third strategy was to examine the impact of deletion of AhR and ARNT on CD8 T-cell function.
The data show that AhR and ARNT expression in CD8 T cells is limited to immune activated effector cells and these transcription factors are not expressed in naïve CD8 T cells. There are only low levels of AhR complexes in conventional CD8 positive cytotoxic T cells. To investigate AhR function at the single cell level we developed a mouse model to fate mark cells that have activated AhR signaling. In this model a mouse expressing Cre recombinase ‘knocked in’ to the CYP1A locus (CYP1A1Cre+/-) was backcrossed to the R26REYFP reporter mouse. In R26REYFP mice, a gene encoding EYFP is knocked into the ubiquitously expressed Rosa26 locus preceded by a loxP flanked stop sequence. CYP1A1 expression is controlled by AhR/ARNT complexes and the concept of our model was that cells that express AhR and ARNT complexes and are triggered with AhR ligands will express Cre recombinase and delete the loxP flanked stop sequence in the R26REYFP reporter locus and hence begin to express YFP.
In vitro experiments demonstrated the validity of this AhR reporter model. The in vitro data reveal that expression of functional AhR/ARNT complexes occurs during Th17 and Tc17 cell differentiation but only a very low frequency of cytotoxic T cells activates the AhR. In vivo data found no evidence for AhR activation during T cell development in the thymus but show strong evidence for activation of AhR/ARNT signaling in innate lymphocytes in the gut.
The ARNT transcription factor is highly expressed in cytotoxic T cells. These cells do not express functional AhR complexes, yet we considered that ARNT might play a role in CD8 T cell biology because of its ability to dimerise with the transcription factor Hif-1a. Our studies of T cells lacking ARNT expression revealed that in CD4 T cells the ARNT transcription factor regulates IL-17 and IL-22 production. In CD8 T cells we discovered that Hif-1a/ARNT signaling controls glycolysis in immune activated cells by sustaining expression of glucose transporters and multiple rate limiting glycolytic enzymes. ARNT was not required for CD8 T cell proliferation but was required for immune activated CD8 T cells to normally differentiate to express perforin and granzymes and to acquire the migratory program of effector T cells. Importantly, we discovered that Hif-1a/ARNT signaling is regulated by mTOR (mammalian target of rapamycin) thus revealing a fundamental mechanism linking nutrient sensing and transcriptional control of CD8 T-cell differentiation.
|Date of Award
|Doreen Cantrell (Supervisor)
- Aryl Hydrocarbon receptor
- T cells