Autophagy Increases Occludin Levels to Enhance Intestinal Paracellular Tight Junction Barrier.

Abstract

Background and AimFunctional loss of paracellular tight junction (TJ) barrier of the gut epithelium and mutations in autophagy genes are factors potentiating inflammatory bowel disease (IBD). Previously we showed the role of autophagy in enhancing the TJ barrier via claudin-2 degradation, however, its role in the regulation of the barrier-forming protein occludin remains unknown. Here, we investigate the role of autophagy in the regulation of occludin and its role in inflammation-mediated TJ barrier loss. MethodsPharmacological and genetic tools were used to study the effect of autophagy on occludin levels and localization, and the role of the MAPK pathway. ResultsAutophagy induction using pharmacological activators and nutrient starvation increased total occludin levels in different epithelial cells. Starvation enriched membrane occludin levels and reduced paracellular inulin flux in Caco-2 cells. Starvation-induced TJ barrier enhancement was contingent on the presence of occludin as OCLN-/- nullified its TJ barrier enhancing effect. Autophagy inhibited the constitutive degradation of occludin and protected against inflammation-induced TJ barrier loss. Starvation-induced TJ barrier enhancement was prevented by inhibition of autophagy. Autophagy enhanced the phosphorylation of ERK-1/2. Inhibition of these kinases in Caco-2 cells and human intestinal mucosa inhibited the protective effects of autophagy. In-vivo, autophagy induction by rapamycin increased occludin levels in mouse intestines and protected against LPS and TNF--induced TJ barrier loss. Additionally, acute Atg7 knockout in adult mice decreased intestinal occludin levels, increasing baseline colonic TJ-permeability and exacerbating the effect of DSS-induced colitis. ConclusionOur data suggest a novel role of autophagy in promoting the intestinal TJ barrier by increasing occludin levels in an ERK1/2 MAPK-dependent mechanism. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=151 SRC="FIGDIR/small/487876v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@1e08d0dorg.highwire.dtl.DTLVardef@14c65f5org.highwire.dtl.DTLVardef@b9e393org.highwire.dtl.DTLVardef@bcd65_HPS_FORMAT_FIGEXP M_FIG C_FIG