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 bo...

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Bibliographic Details
Published in:Molecular biology of the cell 2019-06, Vol.30 (12), p.1406-1424
Main Authors: Van Itallie, Christina M, Lidman, Karin Fredriksson, Tietgens, Amber Jean, Anderson, James Melvin
Format: Article
Language:English
Subjects:
Animals
Calcium - metabolism
Cell Membrane - metabolism
Cell Movement
Claudins - biosynthesis
Dogs
Gene Editing
Half-Life
Kinetics
Madin Darby Canine Kidney Cells
Models, Biological
Occludin - biosynthesis
Tight Junctions - metabolism
Time Factors
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Summary: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. We find that claudins are first delivered to the lateral membrane and, over time, enter the junction strand network from the basal side; this is followed by slow replacement of older claudins in the strands. In contrast, even at early times, newly synthesized occludin is found throughout the network. Taking the results together with our previous documentation of the mechanism for claudin strand assembly in a fibroblast model, we speculate that newly synthesized claudins are added at strand breaks and free ends; these are most common in the basalmost edge of the junction. In contrast, occludin can be added directly within the strand network. We further demonstrate that claudin trafficking and half-life depend on carboxy-terminal sequences and that different claudins compete for tight junction localization.
ISSN:1059-1524
1939-4586
DOI:10.1091/mbc.E19-01-0008