Identification of a novel mechanism of Th2 polarization in mouse CD4+ T cells through nuclear factor erythroid 2-like 2 (Nrf2) regulation

Abstract

Abstract The nuclear factor erythroid 2-like 2 (Nrf2) transcription factor regulates a battery of detoxification and antioxidant genes following activation by cell stress. The primary role of Nrf2 in immune function was thought to occur as an indirect consequence of oxidative stress. We have previously demonstrated an unknown immunomodulatory role of Nrf2, where activation by the common food preservative, tBHQ, induced Nrf2-dependent skewing of CD4+ T cells towards a Th2 phenotype. Polarization resulted in increased production of Th2 cytokines and DNA binding of the master regulator of Th2 differentiation, GATA3. To date, the molecular mechanism(s) responsible for increased GATA3 activity and subsequent Th2 cytokine protein production are unknown and elucidation of this mechanism(s) is the study objective. Primary mouse CD4+ T cells (WT and Nrf2-null) were treated with tBHQ and GATA3 activity was analyzed. Additionally, Nrf2 binding to putative AREs within an intron and the promoter of the GATA3 gene was investigated in the Th2 clone, D10 cells. Results from these studies show GATA3 gene expression and protein production increased after tBHQ treatment in WT, but not Nrf2-null, mice. In addition, Nrf2 binding to putative AREs in the GATA3 gene was demonstrated in D10 cells by ChIP analysis. The functionality of these putative AREs was established by luciferase assay. Together, results of these experiments suggest that GATA3 is upregulated through a direct mechanism involving Nrf2 at the transcriptional level. Overall, these studies provide strong evidence for the role of Nrf2 in a novel mechanism of Th2 polarization of CD4+ T cells, which could have potential implications in Th2 mediated allergic disorders.