Molecular Determinants of the Interaction between Clostridium perfringens Enterotoxin Fragments and Claudin-3

Winkler, Lars; Gehring, Claudia; Wenzel, A.; Müller, Sebastian; Piehl, Christian; Krause, Gerd; Blasig, Ingolf E.; Piontek, Jörg

doi:10.1074/jbc.m109.008623

Cited by 106 publications

(168 citation statements)

References 40 publications

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“…50 The wtCldn3 was previously shown to have strong homology to the human claudin-3 and to specifically bind CPE, whereas the mutCldn3 variant harbors a mutation in the second extracellular loop (for Asn148: N148D; Leu150: L150A), which prevents CPE binding. The mutCldn3-expressing clone served as additional control to further support specificity of toxicity after CPE gene transfer.…”

Section: Transfection Of Human Tumor Cell Linesmentioning

confidence: 99%

Novel Clostridium perfringens enterotoxin suicide gene therapy for selective treatment of claudin-3- and -4-overexpressing tumors

et al. 2011

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Bacterial toxins are known to be effective for cancer therapy. Clostridium perfringens enterotoxin (CPE) is produced by the bacterial Clostridium type A strain. The transmembrane proteins claudin-3 and -4, often overexpressed in numerous human epithelial tumors (for example, colon, breast, pancreas, prostate and ovarian), are the targeted receptors for CPE. CPE binding to them triggers formation of membrane pore complexes leading to rapid cell death. In this study, we aimed at selective tumor cell killing by CPE gene transfer. We generated expression vectors bearing the bacterial wild-type CPE cDNA (wtCPE) or translationoptimized CPE (optCPE) cDNA for in vitro and in vivo gene therapy of claudin-3-and -4-overexpressing tumors. The CPE expression analysis at messenger RNA and protein level revealed more efficient expression of optCPE compared with wtCPE. Expression of optCPE showed rapid cytotoxic activity, hightened by CPE release as bystander effect. Cytotoxicity of up to 100% was observed 72 h after gene transfer and is restricted to claudin-3-and -4-expressing tumor lines. MCF-7 and HCT116 cells with high claudin-4 expression showed dramatic sensitivity toward CPE toxicity. The claudin-negative melanoma line SKMel-5, however, was insensitive toward CPE gene transfer. The non-viral intratumoral in vivo gene transfer of optCPE led to reduced tumor growth in MCF-7 and HCT116 tumor-bearing mice compared with the vector-transfected control groups. This novel approach demonstrates that CPE gene transfer can be employed for a targeted suicide gene therapy of claudin-3-and -4-overexpressing tumors, leading to the rapid and efficient tumor cell killing in vitro and in vivo.

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Section: Transfection Of Human Tumor Cell Linesmentioning

confidence: 99%

Novel Clostridium perfringens enterotoxin suicide gene therapy for selective treatment of claudin-3- and -4-overexpressing tumors

et al. 2011

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“…Cell culture, transfection and immunocytochemistry HEK-293 (Piontek et al, 2011), MDCK-II (Piehl et al, 2010) and Caco-2 (Winkler et al, 2009) cells were cultured in Dulbecco's modified Eagle's medium (DMEM, Invitrogen, Darmstadt, Germany) containing 10% (v/v) fetal calf serum, 100 units/ ml penicillin, 100 mg/ml streptomycin, and 1% L-alanyl-L-glutamine (Invitrogen, Darmstadt, Germany). Transient mono-, co-and triple-transfections were performed using polyethylenimin (PEI) according to the supplier's recommendations (Polysciences, Eppelheim, Germany).…”

Section: Methodsmentioning

confidence: 99%

In tight junctions, claudins regulate the interactions between occludin, tricellulin and marvelD3, which, inversely, modulate claudin oligomerization

Cording

Berg

Käding

et al. 2013

Journal of Cell Science

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158

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SummaryTight junctions seal the paracellular cleft of epithelia and endothelia, form vital barriers between tissue compartments and consist of tight-junction-associated marvel proteins (TAMPs) and claudins. The function of TAMPs and the interaction with claudins are not understood. We therefore investigated the binding between the TAMPs occludin, tricellulin, and marvelD3 and their interaction with claudins in living tight-junction-free human embryonic kidney-293 cells. In contrast to claudins and occludin, tricellulin and marvelD3 showed no enrichment at cell-cell contacts indicating lack of homophilic trans-interaction between two opposing cell membranes. However, occludin, marvelD3 and tricellulin exhibited homophilic cis-interactions, along one plasma membrane, as measured by fluorescence resonance energy transfer. MarvelD3 also cis-interacted with occludin and tricellulin heterophilically. Classic claudins, such as claudin-1 to -5 may show cis-oligomerization with TAMPs, whereas the non-classic claudin-11 did not. Claudin-1 and -5 improved enrichment of occludin and tricellulin at cell-cell contacts. The low mobile claudin-1 reduced the membrane mobility of the highly mobile occludin and tricellulin, as studied by fluorescence recovery after photobleaching. Co-transfection of claudin-1 with TAMPs led to changes of the tight junction strand network of this claudin to a more physiological morphology, depicted by freezefracture electron microscopy. The results demonstrate multilateral interactions between the tight junction proteins, in which claudins determine the function of TAMPs and vice versa, and provide deeper insights into the tight junction assembly.

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“…They dissociate TJ's proteins occludin, ZO-1 and ZO-2, thus increasing intestinal epithelium permeability (Voth and Ballard 2005). The bifunctional enterotoxin CPE of Clostridium perfringens, which can cause diarrhea, forms pores in the host cell plasma membrane, and binds claudin-3 and claudin-4 with high affinity (Sonoda et al 1999;Winkler et al 2009;Robertson et al 2010), inducing their degradation, a decrease in transepithelial resistance and disruption of TJs. The role of claudins in the barrier function of TJs was discovered via these studies (Sonoda et al 1999).…”

Section: Disruption Of Tight Junctions By Enteropathogensmentioning

confidence: 99%

Concepts and Mechanisms: Crossing Host Barriers

Doran

Banerjee

Disson

et al. 2013

Cold Spring Harbor Perspectives in Medicine

111

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Molecular Determinants of the Interaction between Clostridium perfringens Enterotoxin Fragments and Claudin-3

Cited by 106 publications

References 40 publications

Novel Clostridium perfringens enterotoxin suicide gene therapy for selective treatment of claudin-3- and -4-overexpressing tumors

Novel Clostridium perfringens enterotoxin suicide gene therapy for selective treatment of claudin-3- and -4-overexpressing tumors

In tight junctions, claudins regulate the interactions between occludin, tricellulin and marvelD3, which, inversely, modulate claudin oligomerization

Concepts and Mechanisms: Crossing Host Barriers

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