Newly synthesized claudins but not occludin are added to the basal side of the tight junction.

A network of claudin strands creates continuous cell-cell contacts to form the intercellular tight junction barrier; a second protein, occludin, is associated along these strands. The physiological barrier remains stable despite protein turnover, which involves removal and replacement of claudins both in the steady state and during junction remodeling. Here we use a pulse-block-pulse labeling protocol with fluorescent ligands to label SNAP/CLIP-tags fused to claudins and occludin to identify their spatial trafficking pathways and kinetics in Madin-Darby canine kidney monolayers.

MARCKS-related protein regulates cytoskeletal organization at cell-cell and cell-substrate contacts in epithelial cells.

Treatment of epithelial cells with interferon-γ and TNF-α (IFN/TNF) results in increased paracellular permeability. To identify relevant proteins mediating barrier disruption, we performed proximity-dependent biotinylation (BioID) of occludin and found that tagging of MARCKS-related protein (MRP; also known as MARCKSL1) increased ∼20-fold following IFN/TNF administration. GFP-MRP was focused at the lateral cell membrane and its overexpression potentiated the physiological response of the tight junction barrier to cytokines.

Phosphorylation of tight junction transmembrane proteins: Many sites, much to do.

Phosphorylation is a dynamic post-translational modification that can alter protein structure, localization, protein-protein interactions and stability. All of the identified tight junction transmembrane proteins can be multiply phosphorylated, but only in a few cases are the consequences of phosphorylation at specific sites well characterized. The goal of this review is to highlight some of the best understood examples of phosphorylation changes in the integral membrane tight junction proteins in the context of more general overview of the effects of phosphorylation throughout the proteome.