Interferon-gamma induziert Myosin vermittelte Endozytose von Tight Junction Proteinen

Abstract

Tight junctions (TJ) are critical regulators of epithelial paracellular permeability, which is compromised in intestinal mucosa of patients with inflammatory bowel disease. Intestinal inflammation was modeled in vitro by incubating a model intestinal epithelial cell line, T84 with the pro-inflammatory cytokine, interferon-gamma (IFN-γ). We previously found that IFN-γ disrupts epithelial barrier function in model polarized intestinal epithelial cells in an apoptosis independent manner by inducing endocytosis of transmembrane TJ proteins. These proteins were endocytosed into early and recycling endosomes by a macropinocytosis-like process. In the present study, we investigated the role of the actin/myosin cytoskeleton in regulation of TJ proteins endocytosis. Internalized TJ proteins were observed in large vacuoles coated with F-actin and markers of the apical plasma membrane that resemble the vacuolar apical compartment (VAC). Both depolymerization and stabilization of F-actin did not inhibit TJ protein endocytosis suggesting an actin filament turnover independent machinery. Transmembrane TJ protein endocytosis and formation of VACs were blocked by blebbistatin, an inhibitor of non-muscle myosin II A. Phosphorylated myosin light chain, activated form of myosin II A, colocalized with VACs. We conclude that IFN-γ induces endocytosis of TJ transmembrane proteins, occludin, JAM-A, claudin-1 into a vacuolar apical compartment driven by myosin II A mediated formation of VACs.