TNF alpha promotes coxsackie and adenovirus receptor (CAR) phosphorylation by PKC delta to drive leukocyte transepithelial migration

CAR has been established as the primary docking receptor for members of the coxsackie B virus and adenovirus family. More recently, it has been shown that CAR plays an important role in the maintenance of tight junctions and in trans-epithelial migration of leukocytes. These functions have important implications in the immune responses of the respiratory tract, through maintenance of the epithelial barrier function and modulating leucocyte exposure to pathogens. Using epithelial cell lines with mutated versions of the CAR cytoplasmic tail this study has demonstrated that phosphorylation of the cytoplasmic component of CAR promoted transmigration of leukocytes across an epithelial layer. Furthermore, using imaging and western techniques, it was shown that this phosphorylation was triggered by TNF-alpha during inflammation. Using knockdown techniques and chemical inhibition a mechanistic pathway for this phosphorylation effect has been identified. It was shown to occur via stimulation of the enzyme PI-3-K leading to activation and phosphorylation of a PKC delta-dependent pathway. This process was also found to only occur when CAR was homo-dimerised at tight junctions. These events are not confined to immortalised cell lines and have also been demonstrated to be relevant in-vivo. The cytoplasmic tail of CAR was shown to be phosphorylated at cell junctions between epithelial cells of the small airways in both acute (TNF driven) and chronic (ovalbumin driven) inflammatory mouse models. This work identifies a novel mechanism by which the cytoplasmic tail of CAR is involved in the immune response of the respiratory epithelium.