Defining the Functions and Regulation of Clostridium difficile Toxins A and B Enzymatic Activities in Cellular Intoxication

Abstract

Clostridium difficile is a common nosocomial infection that encompasses a range of clinical symptoms from mild diarrhea to complicated pseudomembranous and fulminant colitis. C. difficile secretes two large toxins, TcdA and TcdB, which mediate disease. Understanding the mechanisms by which TcdA and TcdB cause disease is of the utmost importance in designing new, effective, and specific therapies. The toxins are known to bind and enter cells through receptor-mediated endocytosis. Upon exposure to the low pH of the mature endosome, the N-terminal portion of the toxin is delivered to the cytosol and the glucosyltransferase domain is released by an autoprotease cleavage event. The glucosyltransferase domain then targets and modifies small GTPases, resulting in significant cytoskeletal rearrangements. The data presented here offer insights into the mechanisms by which TcdA and TcdB cause cell death and autoprocessing activity is regulated. We demonstrate that TcdA induces a glucuosyltranferase-dependent apoptotic cell death while TcdB causes a concentration-dependent bimodal cell death. At higher concentrations, TcdB induces an enzyme-independent necrotic cell death, and at lower concentrations, TcdB induces an enzyme-dependent apoptotic cell death. We also discovered that zinc is a novel regulator of autoprocessing activity. These data advance our understanding of the functions of C. difficile toxins and provide new targets for drug discovery in the treatment of C. difficile infection.