Cytotoxicity of Clostridium difficile toxins A and B requires an active and functional SREBP‐2 pathway

Clostridium difficile is associated with antibiotic‐associated diarrhea and pseudomembranous colitis in humans. Its 2 major toxins, toxins A and B, enter host cells and inactivate GTPases of the Ras homologue/rat sarcoma family by glucosylation. Pore formation of the toxins in the endosomal membrane enables the translocation of their glucosyltransferase domain into the cytosol, and membrane cholesterol is crucial for this process. Here, we asked whether the activity of the sterol regulatory element–binding protein 2 (SREBP‐2) pathway, which regulates the cholesterol content in membranes, affects the susceptibility of target cells toward toxins A and B. We show that the SREBP‐2 pathway is crucial for the intoxication process of toxins A and B by using pharmacological inhibitors (PF‐429242, 25‐hydroxycholesterol) and cells that are specifically deficient in SREBP‐2 pathway signaling. SREBP‐2 pathway inhibition disturbed the cholesterol‐dependent pore formation of toxin B in cellular membranes. Preincubation with the cholesterol‐lowering drug simvastatin protected cells from toxin B intoxication. Inhibition of the SREBP‐2 pathway was without effect when the enzyme portion of toxin B was introduced into target cells via the cell delivery property of anthrax protective antigen. Taken together, these findings allowed us to identify the SREBP‐2 pathway as a suitable target for the development of antitoxin therapeutics against C. difficile toxins A and B.—Papatheodorou, P., Song, S., López‐Ureña, D., Witte, A., Marques, F., Ost, G. S., Schorch, B., Chaves‐Olarte, E., Aktories, K. Cytotoxicity of Clostridium difficile toxins A and B requires an active and functional SREBP‐2 pathway. FASEB J. 33, 4883–4892 (2019). www.fasebj.org