Clostridium difficile is the leading cause of hospital-acquired
diarrhea in the United States. The two main
virulence factors of C. difficile are the large toxins, TcdA and
TcdB, which enter colonic epithelial cells and cause fluid secretion,
inflammation, and cell death. Using a gene-trap insertional mutagenesis screen, we identified
poliovirus receptor-like 3 (PVRL3) as a cellular factor necessary for
TcdB-mediated cytotoxicity. Disruption of PVRL3 expression by gene-trap mutagenesis,
shRNA, or CRISPR/Cas9 mutagenesis resulted in resistance of cells to
TcdB. Complementation of the gene-trap or CRISPR mutants with PVRL3 resulted in restoration of
TcdB-mediated cell death. Purified PVRL3 ectodomain bound to
TcdB by pull-down. Pretreatment of cells with a
monoclonal antibody against PVRL3 or prebinding
TcdB to PVRL3 ectodomain also inhibited cytotoxicity in cell culture. The receptor is highly expressed on the surface epithelium of the human colon and was observed to colocalize with
TcdB in both an explant model and in tissue from a patient with
pseudomembranous colitis. These data suggest PVRL3 is a physiologically relevant binding partner that can serve as a target for the prevention of
TcdB-induced cytotoxicity in C. difficile
infection.