Endocytosis of epithelial apical junctional proteins by a clathrin-mediated pathway into a unique storage compartment.
The adherens junction (AJ) and tight junction (TJ) are key regulators of epithelial polarity and barrier function. Loss of epithelial phenotype is accompanied by endocytosis of AJs and TJs via unknown mechanisms. Using a model of calcium depletion, we defined the pathway of internalization of AJ and TJ proteins (E-cadherin, p120 and beta-catenins, occludin, JAM-1, claudins 1 and 4, and ZO-1) in T84 epithelial cells. Proteinase protection assay and immunocytochemistry revealed orchestrated internalization of AJs and TJs into a subapical cytoplasmic compartment. Disruption of caveolae/lipid rafts did not prevent endocytosis, nor did caveolin-1 colocalize with internalized junctional proteins. Furthermore, AJ and TJ proteins did not colocalize with the macropinocytosis marker dextran. Inhibitors of clathrin-mediated endocytosis blocked internalization of AJs and TJs, and junctional proteins colocalized with clathrin and alpha-adaptin. AJ and TJ proteins were observed to enter early endosomes followed by movement to organelles that stained with syntaxin-4 but not with markers of late and recycling endosomes, lysosomes, or Golgi. These results indicate that endocytosis of junctional proteins is a clathrin-mediated process leading into a unique storage compartment. Such mechanisms may mediate the disruption of intercellular contacts during normal tissue remodeling and in pathology.
Ivanov
Andrei I
AI
Epithelial Pathobiology Research Unit, Department of Pathology and Laboratory Medicine, Emory University, Atlanta, Georgia 30322, USA. aiivano@emory.edu
Nusrat
Asma
A
Parkos
Charles A
CA
D033965:Adaptor Protein Complex alpha Subunits / Q000378:metabolism; D022005:Adherens Junctions / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D002118:Calcium / Q000494:pharmacology; D051242:Caveolin 1; D022461:Caveolins / Q000378:metabolism / Q000502:physiology; D016764:Cell Polarity / Q000502:physiology; D002478:Cells, Cultured; D002966:Clathrin / Q000378:metabolism / Q000502:physiology; D003598:Cytoskeletal Proteins / Q000378:metabolism / Q000502:physiology; D004705:Endocytosis / Q000502:physiology; D011992:Endosomes / Q000378:metabolism / Q000502:physiology; D004847:Epithelial Cells / Q000378:metabolism* / Q000502:physiology; D006056:Golgi Apparatus / Q000378:metabolism / Q000502:physiology; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000502:physiology; D021962:Membrane Microdomains / Q000378:metabolism; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism* / Q000502:physiology; D008856:Microscopy, Fluorescence; D050765:Qa-SNARE Proteins; D019108:Tight Junctions / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D015534:Trans-Activators / Q000378:metabolism / Q000502:physiology; D051176:beta Catenin
D033965:Adaptor Protein Complex alpha Subunits; C495540:CAV1 protein, human; C495270:CTNNB1 protein, human; D051242:Caveolin 1; D022461:Caveolins; D002966:Clathrin; D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; D050765:Qa-SNARE Proteins; D015534:Trans-Activators; D051176:beta Catenin; D002118:Calcium