The small GTPase Rab13 regulates assembly of functional tight junctions in epithelial cells

Junctional complexes such as tight junctions (TJ) and adherens junctions are required for maintaining cell surface asymmetry and polarized transport in epithelial cells. We have shown that Rab13 is recruited to junctional complexes from a cytosolic pool after cell-cell contact formation. In this stu...

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Bibliographic Details
Published in:Molecular biology of the cell 2002-06, Vol.13 (6), p.1819-1831
Main Authors: Marzesco, Anne-Marie, Dunia, Irene, Pandjaitan, Rudy, Recouvreur, Michel, Dauzonne, Daniel, Benedetti, Ennio Lucio, Louvard, Daniel, Zahraoui, Ahmed
Format: Article
Language:English
Subjects:
Amino Acid Substitution
Animals
Cell Line
Dogs
Epithelial Cells - physiology
Epithelial Cells - ultrastructure
Freeze Fracturing
Microscopy, Electron
Mutagenesis, Site-Directed
rab GTP-Binding Proteins - genetics
rab GTP-Binding Proteins - metabolism
Recombinant Fusion Proteins - metabolism
Recombinant Proteins - metabolism
Tight Junctions - physiology
Tight Junctions - ultrastructure
Transfection
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Summary:Junctional complexes such as tight junctions (TJ) and adherens junctions are required for maintaining cell surface asymmetry and polarized transport in epithelial cells. We have shown that Rab13 is recruited to junctional complexes from a cytosolic pool after cell-cell contact formation. In this study, we investigate the role of Rab13 in modulating TJ structure and functions in epithelial MDCK cells. We generate stable MDCK cell lines expressing inactive (T22N mutant) and constitutively active (Q67L mutant) Rab13 as GFP-Rab13 chimeras. Expression of GFP-Rab13Q67L delayed the formation of electrically tight epithelial monolayers as monitored by transepithelial electrical resistance (TER) and induced the leakage of small nonionic tracers from the apical domain. It also disrupted the TJ fence diffusion barrier. Freeze-fracture EM analysis revealed that tight junctional structures did not form a continuous belt but rather a discontinuous series of stranded clusters. Immunofluorescence studies showed that the expression of Rab13Q67L delayed the localization of the TJ transmembrane protein, claudin1, at the cell surface. In contrast, the inactive Rab13T22N mutant did not disrupt TJ functions, TJ strand architecture nor claudin1 localization. Our data revealed that Rab13 plays an important role in regulating both the structure and function of tight junctions.
ISSN:1059-1524
1939-4586
DOI:10.1091/mbc.02-02-0029