Chondroitin sulfate proteoglycan 4 functions as the cellular receptor

for Clostridium difficile toxin B

Yuan P, Zhang H, Cai C, Zhu S, Zhou Y, Yang X, He R, Li C, Guo S, Li S,

Huang T, Perez-Cordon G, Feng H, Wei W.

Cell Res. 2015 Feb;, 25(2): 157-68.

Speaker: Bo-Yang Tsai (蔡博仰)                              Time: 14:10~15:00, Mar. 18, 2015

Commentator: Dr. I-Hsiu Huang (黃一修老師)           Place: Room 601

Abstract:

     Epidemic Clostridium difficile is the leading cause of antibiotic-associated diarrhea globally due mainily to the production ofits toxin A and toxin B. Both toxins are glucosyl-transferases thatinactivateing Rho family GTPase Rac1 and Cdc42 resulting in cytoskeleton disruption and cell death. However, toxin B has been proven to beided more the essential for high virulence.[1] Previous study demonstrated that the endocytic uptake of both toxins is receptor-dependent. Nevertheless Although the receptor of toxin A has been characterized, toxin B receptor is still unknown, except that is different from toxin A receptor.[2] Here, the authors designed performed a functional screening for toxin B receptor, using the lentivirus encoded shRNAmir library to transfect Hela cell and then incubated with toxin B. Next, gDNA isolated from toxin B-resistant cells was sent was amplified the shRNA encoded region for sequencing. Blast analysis pointed indicated that two differentshRNAs targeteding the same gene encoding fored a surface receptor protein CSPG4 and its role in toxin B-induced cytotoxicity was confirm by TALENs-mediated gene knockout in Hela cell. The binding assay also revealed that the amount of toxin B bound to cell surface correlated with the level of CSPG4 expression by both immunoblotting and flow cytometry, suggesting that CSPG4 is critical for toxin B binding and uptake. To confirm the domain-domain interaction between CSPG4 and toxin B, the authors further constructed many truncated forms of both CSPG4 and toxin B for immunoprecipitation and pull-down assay. TAnd the results demonstrated that a direct interaction between N-terminus of CSPG4 and C-terminus of toxin B and the soluble peptide of toxin-binding domain of CSPG4 could protect cells from the cytotoxicity of toxin B. In However, in vivo experiment demonstrated that, the complete loss of CSPG4/NG2 could abolish the toxin B-triggered IL-8 induction without significantly affecting mice viability. In conclusion, the authors report that the first identification of the toxin B functional receptor and reveals a previously unsuspected role for CSPG4.

References:

1.         Lyras, D., et al., Toxin B is essential for virulence of Clostridium difficile. Nature, 2009. 458(7242): p. 1176-9.

2.         Na, X., et al., gp96 is a human colonocyte plasma membrane binding protein for Clostridium difficile toxin A. Infect Immun, 2008. 76(7): p. 2862-71.