Ligand-Independent Regulation of Transforming Growth Factor β1 Expression and Cell Cycle Progression by the Aryl Hydrocarbon Receptor

Abstract

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that mediates the toxic effects of its xenobiotic ligands and acts as an environmental checkpoint during the cell cycle. We expressed stably integrated, Tet-Off-regulated AHR variants in fibroblasts from AHR-null mice to further investigate the AHR role in cell cycle regulation. Ahr+/+ fibroblasts proliferated significantly faster than Ahr-/- fibroblasts did, and exposure to a prototypical AHR ligand or deletion of the ligand-binding domain did not change their proliferation rates, indicating that the AHR function in cell cycle was ligand independent. Growth-promoting genes, such as cyclin and cyclin-dependent kinase genes, were significantly down-regulated in Ahr-/- cells, whereas growth-arresting genes, such as the transforming growth factor beta1 (TGF-beta1) gene, extracellular matrix (ECM)-related genes, and cyclin-dependent kinase inhibitor genes, were up-regulated. Ahr-/- fibroblasts secreted significantly more TGF-beta1 into the culture medium than Ahr+/+ fibroblasts did, and Ahr-/- showed increased levels of activated Smad4 and TGF-beta1 mRNA. Inhibition of TGF-beta1 signaling by overexpression of Smad7 reversed the proliferative and gene expression phenotype of Ahr-/- fibroblasts. Changes in TGF-beta1 mRNA accumulation were due to stabilization resulting from decreased activity of TTP, the tristetraprolin RNA-binding protein responsible for mRNA destabilization through AU-rich motifs. These results show that the Ah receptor possesses interconnected intrinsic cellular functions, such as ECM formation, cell cycle control, and TGF-beta1 regulation, that are independent of activation by either exogenous or endogenous ligands and that may play a crucial role during tumorigenesis.