Formation of tight junction: determinants of homophilic interaction between classic claudins.
Claudins are the critical transmembrane proteins in tight junctions. Claudin-5, for instance, prevents paracellular permeation of small molecules. However, the molecular interaction mechanism is unknown. Hence, the claudin-claudin interaction and tight junction strand formation were investigated using systematic single mutations. Claudin-5 mutants transfected into tight junction-free cells demonstrated that the extracellular loop 2 is involved in strand formation via trans-interaction, but not via polymerization, along the plasma membrane of one cell. Three phenotypes were obtained: the tight junction type (wild-type-like trans- and cis-interaction; the disjunction type (blocked trans-interaction); the intracellular type (disturbed folding). Combining site-directed mutagenesis, live-cell imaging-, electron microscopy-, and molecular modeling data led to an antiparallel homodimer homology model of the loop. These data for the first time explain how two claudins hold onto each other and constrict the paracellular space. The intermolecular interface includes aromatic (F147, Y148, Y158) and hydrophilic (Q156, E159) residues. The aromatic residues form a strong binding core between two loops from opposing cells. Since nearly all these residues are conserved in most claudins, our findings are of general relevance for all classical claudins. On the basis of the data we have established a novel molecular concept for tight junction formation.
Piontek
Jörg
J
Leibniz-Institut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany.
Winkler
Lars
L
Wolburg
Hartwig
H
Müller
Sebastian L
SL
Zuleger
Nikolaj
N
Piehl
Christian
C
Wiesner
Burkhard
B
Krause
Gerd
G
Blasig
Ingolf E
IE
D019943:Amino Acid Substitution; D002460:Cell Line; D062507:Claudin-5; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D007150:Immunohistochemistry; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D016297:Mutagenesis, Site-Directed; D013347:Subcellular Fractions / Q000378:metabolism; D019108:Tight Junctions
C107466:CLDN5 protein, human; D062507:Claudin-5; D008565:Membrane Proteins