66 Bacterial toxins Clostridium perfringens enterotoxin Food poisoning by Clostridium perfringens enterotoxin (CPE) leads to diarrhea symptoms classiWed as “C. perfrin- gens type A food poisoning and antibiotic-associated diar- rhea”. The enterotoxin protein has been elucidated to intrude the cells by interacting with epithelial TJ, including certain claudins. Toxin-induced cytolytic pore-formation is a prerequisite for this and involves residues in the N-termi- nal half of CPE protein, while residues near the C-terminus are required for binding to claudins. In detail, the single 35 kDa CPE polypeptide Wrst causes cellular damage by altering plasma membrane permeability by the formation of an approximately 155 kDa CPE-containing pore complex necessary for the subsequent interaction with TJ proteins via its C-terminal receptor-binding region (McClane 2001). CPE was then perceived to aVect TJ structure and function in a way that permeability properties of the epithelial layer are altered leading to diarrhea (McClane 2001). Further investigation has determined the isoforms of claudins targeted by CPE. Sonoda et al. (1999) were able to show that the COOH-terminal half fragment of CPE (C- CPE) bound eVectively to claudin-3 and claudin-4 in MDCK I cells expressing the claudins-1, -2, -3 and -4. They were further able to show that in the presence of C-CPE, reconstituted TJ strands gradually disintegrated and disappeared from their cell surface in C3L cells. As a consequence, because diverse claudins are overex- pressed on some human cancers, the toxin became interest- ing for targeting chemotherapy in the manner of a Trojan horse (Long et al. 2001; Michl et al. 2001). Recently, van Itallie et al. (2007) aimed to solve the structure of the CPE claudin-binding domain to advance its therapeutic applications. The structure was shown to be a nine-strand beta sandwich with previously unappreciated similarity to the receptor-binding domains of several other toxins of spore-forming bacteria, giving strong evidence for the presence of a common ancestor for several receptor- binding domains of bacterial toxins (Van Itallie et al. 2007). The very recent elucidation of the structure of a 14- kDa fragment containing residues 194 to the native COOH terminus at position 319 by X-ray diVraction to a resolution below 2 Å showed that the structure is a nine-strand sand- wich with previously unappreciated similarity to the recep- tor-binding domains of several other toxins of spore- forming bacteria, i.e., the collagen-binding domain of ColG from Clostridium histolyticum and the large Cry family of toxins (including Cry4Ba) of Bacillus thuringiensis (Van Itallie et al. 2008). The authors speculate that the claudin-4 binding site is on a large surface loop between strands 8 and 9 or includes both of these strands. They further were 123 Histochem Cell Biol (2008) 130:55–70 able to clarify that the sequence crystallized binds to puri- Wed human claudin-4 with a 1:1 stoichiometry. The binding aYnity was determined to be in the submicromolar range similar to that observed for binding of native toxin to cells (Van Itallie et al. 2008). These results could now provide a structural framework to advance therapeutic applications of the toxin. Conclusion and perspectives The discovery of the existence of occludin- and claudin- based TJs constituting a permeability barrier in epithelial and endothelial cells has paved the way for the elucidation of the molecular background of diverse human diseases the pathogenesis of which had been unclear before. It became clear that the presence of TJs is indispensable for tissue compartmentalisation and cellular homeostasis. Distur- bances in TJ function are reXected in many diseases, but on the other hand, awareness of their involvement has facili- tated therapy enormously. For instance, antiinXammatory or antimetastatic drugs could be developed based on their ability to repair broken barrier function. These strategies have become indispensable in the treatment of disease like MS, inXammatory bowel disease or diabetic retinopathy. Following the strategy of pathogenic agents, which use TJ proteins as a docking station to invade cells, even strategies for drug delivery techniques could be implemented. Future attempts will have to address the role of occludin and the exact contribution of the diverse claudins to the formation of ion selective pores, which are still open questions. A more thorough understanding will greatly facilitate diagno- sis and the development of speciWc treatment regimens for diseases originating from impaired TJ function. Acknowledgments This work was supported by grant SFB688/ TPA5 from Deutsche Forschungsgemeinschaft DFG to CF. The author is especially thankful toDetlev Drenckhahn for constructive sugges- tions and critical reading of the manuscript. The author is grateful to Kinga Blecharz for editing expertise and Malgorzata Burek for eY- cient and pleasant help in collection of material. References Abbott NJ, Ronnback L, Hansson E (2006) Astrocyte-endothelial interactions at the blood-brain barrier. Review. Nat Rev Neurosci 7:41–53 Achler C, Filmer D, Merte C, Drenckhahn D (1989) Role of microtu- bules in polarized delivery of apical membrane proteins to the brush border of the intestinal epithelium. J Cell Biol 109:179–189 Ahdieh M, Vandenbos T, Youakim A (2001) Lung epithelial barrier function and wound healing are decreased by IL-4 and IL-13 and enhanced by IFN-gamma. 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