Functional analysis of clostridium sordellii lethal and hemorrhagic toxins
Craven, Ryan Eric
Clostridium sordellii infections cause gangrene and edema in humans and gastrointestinal infections in livestock. The two principle virulence factors, TcsH and TcsL, are highly homologous to C. difficile TcdA and TcdB, respectively. Experiments to compare the effects of TcsH and TcsL to TcdA and TcdB show that TcsH induces necrotic cell death similarly to TcdB while TcsL induces apoptotic cell death. Further work focuses on TcsL, which has two enzymatic domains, an N-terminal glucosyltransferase domain (GTD) and an autoprocessing domain responsible for release of the GTD within the cell. The GTD can then use its N-terminal membrane localization domain (MLD) for orientation on membranes and modification of GTPases. The use of conditionally immortalized murine pulmonary microvascular endothelial cells provides a model for the study of TcsL functional activities. Point mutations that disrupt the glucosyltransferase, autoprocessing, or membrane localization activities are introduced into a recombinant version of TcsL, and the activities of these mutants are compared to those of wild-type toxin. All mutants are defective or impaired in cytotoxicity but differ in their modification of Rac1 and Ras. The data suggest a model where differences in GTPase localization dictate cellular responses to intoxication and highlight the importance of autoprocessing in the function of TcsL.