Mechanism of the paracellular molecular transport

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To understand the tight junction (TJ) as the apparatus for molecular transfer and physiological barrier at the paracellular space, we analyzed function of claudins composing TJ and the mode of action by which bacterial virulence factors affect the paracellular barrier function of TJ.1. Functional analyses of claudins.(1) We generated claudin-1-deficient and claudin-5-deficient mice, respectively, and found that claudin-1 constitutes TJ that was first demonstrated in the epidermis, and claudin-5 essentially forms TJ at the blood-brain barrier.(2) We analyzed dynamic behavior of the TJ strand by using Eph4 epithelial cells expressing GFP-tagged claudin. It was found that the TJ strands retain their continuities during their remodeling accompanied by the cell movement When the TJ strands shortened, the superfluous claudins were endocytosed into one of the adjacent cells. This dynamic behavior is considered to underlie the constitutive barrier function of TJ.2. Analyses for mechanisms by which bacterial virulence factors affect the barrier function of TJ(1) We examined how enteropathogenic Escherichia coli (EPEC) reduces the paracellular barrier function in the epithelial cell model, and found that Map, one of EPEC virulence factors, is responsible for it Moreover, the intimate association between EPEC and epithelial cells was found to trigger the secretion of (a) virulence factor(s) other than Map, which are essential for the barrier-disrupting activity of EPEC.(2) We analyzed the binding nature between claudins and Clostridium perfiivens enterotoxin (CPE), which is known to specifically recognize claudins as receptors. The results demonstrated that CPE binds to claudin 6,7,8, and 14 besides claudin 3 and 4, which had been reported so far, and that CPE recognizes the second extracellular loop of the claudins. Both in vitro and in vivo experiments revealed that a claudin-binding fragment of CPE opened the paracellular pathways of epithelial cells.