Fig. 2

The TJ opening Claudin5a^Y148A mutant increases TJ permeability. (A) Schematic representation of the four-transmembrane pass TJ protein Claudin5a (Cldn5a). The two extracellular loops (ECL), facing the paracellular space, are involved in claudin-claudin trans interactions. The position of an aromatic residue within ECL2, which is conserved between all classic claudins (17) and which is essential for trans interactions, is highlighted in red. ECL2, which is conserved between all classic claudins (17) and which is essential for trans interactions. (B) Stable transfection of murine Cldn5^wt reduces the paracellular permeability of fluorescein (filter culture) compared to the vector control, whereas transfection of the mutant Cldn5^Y148A causes an increase of fluorescein permeability. Data represent mean ± SEM, n ≥ 10; *, P < 0.05, **, P < 0.01. (C) Stable transfection of murine Cldn5^wt and Cldn5^Y148A into MDCK-II cells. The Cldn5 variants are strongly enriched at cell-cell contacts as detected by immunocytochemistry against the N-terminal FLAG-tag. (Scale bars: 10 μm.) (D) Injection of mRNA encoding zebrafish Cldn5a^wt or Cldn5a^Y148A into cldn5a morphants results in the correct localization of the proteins at the ventricular surface of the 30 hpf neural tube as detected by confocal microscopy of sectioned immunohistochemical stainings. (Scale bar: 50 μm.)