Functional defects in Clostridium difficile TcdB toxin uptake identify CSPG4 receptor-binding determinants

Area of Science:

• Microbiology
• Cell Biology
• Toxicology

Background:

• Clostridium difficile produces toxins TcdA and TcdB, crucial for disease.
• TcdB is a primary cause of C. difficile-related human illness.
• TcdB utilizes host cell receptors for entry, with CSPG4, NECTIN3, and FZD1/2/7 identified.

Purpose of the Study:

• To identify specific binding sites of TcdB on host cell receptors.
• To investigate the role of the combined repetitive oligopeptide (CROP) domain in TcdB receptor binding.
• To understand the mechanism of TcdB-mediated cell intoxication.

Main Methods:

• Screening TcdB for essential intoxication sites.
• Utilizing C-terminal truncations to map binding regions.
• Testing TcdB binding and toxicity on CSPG4-expressing cells.
• Assessing the impact of anti-TcdB antibody bezlotoxumab on CSPG4 binding.

Main Results:

• A region at the translocation and CROP domain junction is implicated in CSPG4 binding.
• The CSPG4 binding site extends into the CROP domain, requiring three short repeats.
• Bezlotoxumab binding partially overlaps the CSPG4 binding site, inhibiting binding.
• The CROP domain plays a role in TcdB receptor binding.

Conclusions:

• The CSPG4 binding site on TcdB has been localized to the translocation-CROP domain junction and within the CROP domain.
• This study demonstrates a novel role for the TcdB CROP domain in host cell receptor binding.
• Findings clarify TcdB pathogenesis and provide insights for therapeutic strategies targeting TcdB-host interactions.