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Caserta, Justin Angelo (2010) CONTRIBUTION OF COMPLEX FORMATION IN THE IN VITRO AND IN VIVO ACTION OF CLOSTRIDIUM PERFRINGENS ENTEROTOXIN. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Clostridium perfringens enterotoxin (CPE) is a pore-forming toxin that is responsible for causing the symptoms of type A food poisoning, a leading cause of bacterial foodborne illness in the US. CPE-induced pore formation on intestinal epithelial cells results in ion permeability alterations leading to Ca2+ influx and activation of cell death pathways. Upon binding to its receptor, certain claudins, the interactions between CPE and the target membrane result in the formation of a series of toxin complexes (CH-1 and CH-2) that represent the formation of the functional CPE pore. Many bacterial toxins, particularly, pore-forming toxins, hijack cholesterol-rich lipid raft domains in the target cell membrane to aid in their virulence. Lipid rafts serve as platforms to cluster receptor proteins to allow for more efficient binding and oligomerization. Due to the pore-forming activity of CPE, we wished to determine if membrane rafts play a role in the mechanism of action of CPE. Interestingly, CPE was found to be a novel pore-forming toxin that does not require raft domains for its action in that CPE complexes do not form within lipid rafts and cholesterol depletion had no effect on CPE-induced cytotoxicity. These findings illustrate the unique interactions between CPE and target cells. Despite recent research findings indicating the presence of claudins in the various CPE complexes, these intricate interactions have not been fully elucidated, and the exact composition of the toxin complexes is unknown. Therefore, the research presented here describes the development of a two-step method of electroelution/immunoprecipitation that allows for the isolation and purification of the CPE complexes for compositional analysis by mass spectrometry. Finally, a mouse model has been developed and characterized to show that the molecular interactions that occur in cell culture models, such as complex formation and inflammatory cell death, also occur in vivo. Furthermore, the mouse model mimics the lethality that is occasionally seen in humans that suffer from type A food poisoning-related deaths.


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Item Type: University of Pittsburgh ETD
Status: Unpublished
CreatorsEmailPitt UsernameORCID
Caserta, Justin Angelojac86@pitt.eduJAC86
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Committee ChairMcClane, Bruce Abamcc@pitt.eduBAMCC
Committee MemberDay, Billybday@pitt.eduBDAY
Committee MemberParniak, Mikemap167@pitt.eduMAP167
Committee MemberDeLuca, Nealndeluca@pitt.eduNDELUCA
Committee MemberSalter, Russellrds@pitt.eduRDS
Date: 28 July 2010
Date Type: Completion
Defense Date: 9 July 2010
Approval Date: 28 July 2010
Submission Date: 27 July 2010
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Institution: University of Pittsburgh
Schools and Programs: School of Medicine > Molecular Virology and Microbiology
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: Clostridia; enterotoxin; microbiology; pore-forming toxin; raft
Other ID:, etd-07272010-114552
Date Deposited: 10 Nov 2011 19:54
Last Modified: 15 Nov 2016 13:47


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