Functional analyses of epidemic Clostridioides difficile toxin B variants reveal their divergence in utilizing receptors and inducing pathology

Zhenrui Pan; Yuanyuan Zhang; Jianhua Luo; Danyang Li; Yao Zhou; Liuqing He; Qi Yang; Min Dong; Liang Tao

doi:10.1371/journal.ppat.1009197

Functional analyses of epidemic Clostridioides difficile toxin B variants reveal their divergence in utilizing receptors and inducing pathology

PLoS Pathog. 2021 Jan 28;17(1):e1009197. doi: 10.1371/journal.ppat.1009197. eCollection 2021 Jan.

Authors

Zhenrui Pan^{1

2

3}, Yuanyuan Zhang^{1

2

3}, Jianhua Luo^{1

2

3}, Danyang Li^{1

2

3}, Yao Zhou^{1

2

3}, Liuqing He^{1

2

3}, Qi Yang^{1

2

3}, Min Dong^{4

5}, Liang Tao^{1

2

3}

Affiliations

¹ Key Laboratory of Structural Biology of Zhejiang Province, School of Life Sciences, Westlake University, Hangzhou, China.
² Center for Infectious Disease Research, Westlake Laboratory of Life Sciences and Biomedicine, Hangzhou, China.
³ Institute of Basic Medical Sciences, Westlake Institute for Advanced Study, Hangzhou, China.
⁴ Department of Urology, Boston Children's Hospital, Boston, Massechusetts, United States of America.
⁵ Department of Surgery and Department of Microbiology, Harvard Medical School, Boston, Massechusetts, United States of America.

Abstract

Clostridioides difficile toxin B (TcdB) is a key virulence factor that causes C. difficile associated diseases (CDAD) including diarrhea and pseudomembranous colitis. TcdB can be divided into multiple subtypes/variants based on their sequence variations, of which four (TcdB1-4) are dominant types found in major epidemic isolates. Here, we find that these variants are highly diverse in their receptor preference: TcdB1 uses two known receptors CSPG4 and Frizzled (FZD) proteins, TcdB2 selectively uses CSPG4, TcdB3 prefers to use FZDs, whereas TcdB4 uses neither CSPG4 nor FZDs. By creating chimeric toxins and systematically switching residues between TcdB1 and TcdB3, we determine that regions in the N-terminal cysteine protease domain (CPD) are involved in CSPG4-recognition. We further evaluate the pathological effects induced by TcdB1-4 with a mouse intrarectal installation model. TcdB1 leads to the most severe overall symptoms, followed by TcdB2 and TcdB3. When comparing the TcdB2 and TcdB3, TcdB2 causes stronger oedema while TcdB3 induces severer inflammatory cell infiltration. These findings together demonstrate divergence in the receptor preference and further lead to colonic pathology for predominant TcdB subtypes.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Bacterial Proteins / genetics*
Bacterial Proteins / metabolism*
Bacterial Toxins / genetics*
Bacterial Toxins / metabolism*
Chondroitin Sulfate Proteoglycans / genetics
Chondroitin Sulfate Proteoglycans / metabolism*
Clostridioides difficile / genetics
Clostridioides difficile / metabolism*
Clostridium Infections / genetics
Clostridium Infections / metabolism
Clostridium Infections / microbiology
Clostridium Infections / pathology*
Epidemics
Female
Frizzled Receptors / genetics
Frizzled Receptors / metabolism*
HeLa Cells
Humans
Mice
Mice, Inbred BALB C
Mutation*
Virulence Factors / genetics
Virulence Factors / metabolism

Substances

Bacterial Proteins
Bacterial Toxins
Chondroitin Sulfate Proteoglycans
Frizzled Receptors
Virulence Factors
toxB protein, Clostridium difficile

Grants and funding

This study was partially supported by the National Natural Science Foundation of China (Grant no. 31970129 and Grant no. 31800128 to L.T.). L.T. also acknowledges support by the Zhejiang Provincial Natural Science Foundation of China under Grant no. LR20C010001, Westlake Education Foundation (http://www.wefoundation.org.cn/), and Tencent Foundation (https://gongyi.qq.com/jjhgy/). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.