Identification of an epithelial cell receptor responsible for Clostridium difficile TcdB-induced cytotoxicity.

Abstract

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.

• Authors: Michelle E LaFrance, Melissa A Farrow, Ramyavardhanee Chandrasekaran, Jinsong Sheng, Donald H Rubin, D Borden Lacy
• Journal: Proceedings of the National Academy of Sciences of the United States of America (Proc Natl Acad Sci U S A) Vol. 112 Issue 22 Pg. 7073-8 (Jun 02 2015) ISSN: 1091-6490 [Electronic] United States
• PMID: 26038560 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• Antibodies, Monoclonal
• Bacterial Proteins
• Bacterial Toxins
• Cell Adhesion Molecules
• Enterotoxins
• NECTIN3 protein, human
• Nectins
• toxB protein, Clostridium difficile

Topics

• Analysis of Variance
• Antibodies, Monoclonal (metabolism)
• Bacterial Proteins (metabolism, toxicity)
• Bacterial Toxins (metabolism, toxicity)
• Caco-2 Cells
• Cell Adhesion Molecules (genetics, immunology, metabolism)
• Clostridioides difficile (chemistry)
• Colon (metabolism)
• Enterotoxins (metabolism, toxicity)
• Epithelial Cells (metabolism)
• Genetic Complementation Test
• HeLa Cells
• Humans
• Mutagenesis, Insertional
• Nectins