significance
Tight junction form selective osmotic seals that limit paracellular flux. Increased tight junction permeability is associated with intestinal disease. Previous studies have shown that casein kinase 2 (CK2) phosphorylates S408 within an unstructured region of the cytoplasmic tail of the tight junction protein occludin. Furthermore, CK2 inhibition and consequent S408 dephosphorylation reduced paracellular permeability in vitro and in vivo, and attenuated experimental immune-mediated intestinal disease in vivo. Here, we show that S408 phosphorylation enhances intramolecular interactions between the unstructured region and the distal α-helical bundle to restrict binding to the tight junction protein zonula occludens-1 (ZO-1). Conversely, S408 dephosphorylation restricted intramolecular interactions and promoted occludin binding to ZO-1 in vitro.
Abstract
The paracellular channels through which ions and small molecules pass through epithelial cells are controlled by tight junction, a complex network of claudin polymers that form tight seals between adjacent cells. How the nanoscale structure of tight junction networks enables paracellular channels of specific ions or small molecules without compromising barrier function is unclear. Here, we combine super-resolution stimulated emission depletion microscopy, multivariate sorting of super-resolution images, and fluorescence resonance energy transfer in living and fixed cells and tissues to reveal the tight junction formed by mammalian claudins of nanoscale organization. We show that only a fraction of claudins can assemble into characteristic homotypic networks, whereas tight junction formed by multiple claudins display nanoscale organizing principles for mixing, integrating, inducing, separating, and excluding chain assembly. Interestingly, channel-forming claudins are spatially separated from barrier-forming claudins by determinants encoded primarily in their extracellular domains, which are also known to harbor human disease-causing mutations. Electrophysiological analysis of claudin in epithelial cells demonstrates that nanoscale separation of distinct channel-forming claudin enables barrier function to be coupled with specific paracellular ion fluxes across tight junction. Channel-forming claudins are spatially separated from barrier-forming claudins by determinants encoded primarily in their extracellular domains, which are also known to harbor human disease-causing mutations. Electrophysiological analysis of claudin in epithelial cells demonstrates that nanoscale separation of distinct channel-forming claudin enables barrier function to be coupled with specific paracellular ion fluxes across tight junction. Channel-forming claudins are spatially separated from barrier-forming claudins by determinants encoded primarily in their extracellular domains, which are also known to harbor human disease-causing mutations. Electrophysiological analysis of claudin in epithelial cells demonstrates that nanoscale separation of distinct channel-forming claudin enables barrier function to be coupled with specific paracellular ion fluxes across tight junction.
introduce
During development and homeostasis, tissues must not only tightly control the passage of small molecules and ions through transcellular transport mechanisms, but also through paracellular adhesion complexes, including tight junction (TJs) 1 . pass. TJs form apical cell-cell contacts in epithelial cells2 and restrict the passage of pathogens, small molecules, ions and water through very tight paracellular membrane contacts1. Claudin is a transmembrane protein that forms chains about 10 nm thick, interwoven into a TJ network3, and binds to a number of scaffolding proteins and the cytoskeleton4 intracellularly. These chains can act as paracellular diffusion barriers against large and small solutes, For example, when composed of barrier claudins such as claudin-1 (Cldn1) 5, 6 or Cldn3 7 . Furthermore, Cldn2 8 , 9 , 10 , Cldn10a/b 11 , 12 , Cldn15 13 , 14 , 15 and Cldn16 16 , 17 formed size- and charge-selective ion/water channels within the chain.