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Tight junction targeting and intracellular trafficking of occludin in polarized epithelial cells. Am J Physiol Cell Physiol 2007 Nov;293(5):C1717-26

Date

09/15/2007

Pubmed ID

17855770

DOI

10.1152/ajpcell.00309.2007

Scopus ID

2-s2.0-36049047582   26 Citations

Abstract

Occludin, a transmembrane (TM)-spanning protein, is an integral component of the tight junctional (TJ) complexes that regulate epithelial integrity and paracellular barrier function. However, the molecular determinants that dictate occludin targeting and delivery to the TJs remain unclear. Here, using live cell imaging of yellow fluorescent protein-labeled occludin fragments, we resolved the intracellular trafficking of occludin-fusion proteins in polarized Madin-Darby canine kidney and Caco-2 cells to delineate the regions within the occludin polypeptide that are important for occludin targeting to the TJs. Live cell confocal imaging showed that complete or partial truncation of the COOH-terminal tail of the occludin polypeptide did not prevent occludin targeting to the TJs in epithelial cell lines. Progressive truncations into the COOH-terminal tail decreased the efficiency of occludin expression; after the removal of the regions proximal to the fourth transmembrane domain (TM4), the efficiency of expression increased. However, further deletions into the TM4 abolished TJ targeting, which resulted in constructs that were retained intracellularly within the endoplasmic reticulum. The full-length occludin polypeptide trafficked to the cell surface within a heterogenous population of intracellular vesicles that delivered occludin to the plasma membrane in a microtubule- and temperature-dependent manner. In contrast, the steady-state localization of occludin at the cell surface was dependent on intact microfilaments but not microtubules.

Author List

Subramanian VS, Marchant JS, Ye D, Ma TY, Said HM

Author

Jonathan S. Marchant PhD Chair, Professor in the Cell Biology, Neurobiology and Anatomy department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Animals
Caco-2 Cells
Calcium
Cell Polarity
Colchicine
Cytochalasin D
Cytoplasm
Cytoskeleton
Dogs
Endoplasmic Reticulum
Epithelial Cells
Green Fluorescent Proteins
Humans
Intestinal Mucosa
Intestines
Kidney
Magnesium
Membrane Proteins
Microscopy, Confocal
Nocodazole
Occludin
Peptide Fragments
Peptide Mapping
Protein Structure, Tertiary
Protein Transport
Recombinant Fusion Proteins
Temperature
Tight Junctions
Transfection
Transport Vesicles
Tubulin Modulators
jenkins-FCD Prod-482 91ad8a360b6da540234915ea01ff80e38bfdb40a