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Disruption of Barrier Integrity by Salmonella typhimurium Requires Activation of Cdc42 and Rac1 in Epithelial Cells
Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
(English)Manuscript (preprint) (Other academic)
Abstract [en]

Epithelial cells that line the human intestinal mucosa constitute the initial sites of host invasion by bacterial pathogens. A number of bacteria, such as Salmonella and Yersinia, have been shown to disrupt the integrity of the epithelial barrier, although little is known about the mechanisms underlying that effect. We found that polarized MDCK-1 epithelial cells infected with invasive Salmonella typhimurium SLI344 exhibited marked changes in F-actin organization and a rapid decrease in transepithelial electrical resistance (TER). In contrast, infection with isogenic noninvasive mutants (hilA, prgH, and sipC) increased the TER in these cells. Pretreating MDCK-1 cells with the tyrosine kinase inhibitor genistein or the PI-3 kinase inhibitor wortmannin did not affect invasion and subsequent perturbation of the epithelial barrier by S. typhimurium. Instead, the specific geranylgeranyltransferase-1 inhibitor GGTI-298, but not the farnesyltransferase inhibitor FTI-277, clearly reversed the capacity of S. typhimurium to disrupt the epithelial barrier. The best-known substrates for GGTI-298 include Rho family GTPases, Racl and Cdc42. Infection with wild-type S. typhimurium increased the level of activated Racl and Cdc42 and caused these proteins to accumulate apically in MDCK-1 cells. GGTI-298-induced inactivation of Racl and Cdc42 prevented alteration of the tight and adherens junction-associated proteins Z0-1, occludin, and E-cadherin in MDCK-1 cells infected with invasive Salmonella. These results indicate that activation of Racl and Cdc42, but not tyrosine kinase or PI-3 kinase, is essential for disruption of barrier integrity by S. typhimurium in polarized MDCK-1 cells.

Keyword [en]
Salmonella typhimurium, epithelium, tight junctions, Rac1, Cdc42
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:liu:diva-80200OAI: oai:DiVA.org:liu-80200DiVA: diva2:546129
Available from: 2012-08-22 Created: 2012-08-22 Last updated: 2012-08-22Bibliographically approved
In thesis
1. Perturbation of the epithelial barrier by enteric pathogens
Open this publication in new window or tab >>Perturbation of the epithelial barrier by enteric pathogens
2001 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Gastrointestinal infections in humans have been associated with a number of diseased condition, including stomach ulcers, gastroenteritis, Crohn's disease, and rheumatic arthritis. Such infections often cause altered intestinal permeability through perturbation of the tight junctions that hold epithelial cells together. The objective of the present studies was to detennine whether the enteric pathogens Salmonella, Yersinia, and Rotavirus can disrupt the integrity of the epithelial barrier, and, if so, how this is achieved. Another aim was to elucidate regulation of the epithelial batrier in relation to the structure of the cytoskeleton.

To accomplish these goals, we assessed the mechanism of enhanced cytotoxicity of Yersinia YopE and the response to this protein by its target in the epithelial bartier, both of which require contact between the bacteria and the eukaryotic cells. YopK appeared to control Yop effector delivery by regulating the size of the translocation pore, and enhanced translocation was accompanied by decreased transepithelial resistance and disruption of barrier function. We also examined the interaction of Yersinia with polarized MDCK cells to detemrine the target of these bacteria. We found that wild-type Yersinia adhered apically to the tight junction areas, and, in adjacent cells, these contact points displayed ß1 integrins and tight junction proteins that allowed localized invasin-mediated binding and translocation of cytotoxins. Studying signal transduction pathways involved in the disruption of barrier function by Salmonella typhimurium, we found that infection with the wild-type strain increased the level of activated. Rac1 and Cdc42 small G-proteins and caused them to accumulate apically in MDCK cells, and this was prevented by appropriate inhibitors. Activation of these proteins was a prerequisite of disruption of barrier integrity by S. typhimurium. We also considered specific effects of the rota virus non-structural protein NSP4 on the function of tight junctions. NSP4 has been desctibed as the first viral enterotoxin, and we found that incubation of noncontluent MDCK-1 cells with NSP4 prevented development of the permeability barrier, as well as lateral targeting of the tight junction-associated zonula occludence-1 protein.

In conclusion, our results provide strong evidence that the studied pathogens perturb the epithelial barrier by binding to specific cell receptors to deliver cytotoxins (Yesinia); by interfering with cell signaling pathways (Salmonella); and by impairing normal formation of tight junctions (NSP4).

Place, publisher, year, edition, pages
Linköping: Linköpings universitet, 2001. 58 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 702
Keyword
enteric pathogens: signal transduction, barrier function
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-28644 (URN)13800 (Local ID)91-7373-146-3 (ISBN)13800 (Archive number)13800 (OAI)
Public defence
2001-12-03, Berzeliussalen, Universitetssjukhuset, Linköping, 13:00 (Swedish)
Opponent
Supervisors
Available from: 2009-10-09 Created: 2009-10-09 Last updated: 2012-11-22Bibliographically approved

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Tafazoli, FaridehMagnusson, Karl-EricZheng, Liming

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