Impact of clostridial glucosylating toxins on the proteome of colonic cells determined by isotope-coded protein labeling and LC-MALDI

Proteome Science

Research

Nelli Jochim¹  Andreas Pich¹  Ingo Just¹  Ralf Gerhard¹ 

[1] Hannover Medical School, Institute of Toxicology, Carl-Neuberg-Str. 1, D-30625, Hannover, Germany;

关键词: C. difficile;    colonic cells;    ICPL™;    relative quantification;    Toxin A;   

DOI  :  10.1186/1477-5956-9-48

 received in 2011-05-31, accepted in 2011-08-17,  发布年份 2011

来源: Springer

【 摘 要 】

BackgroundThe anaerobe Clostridium difficile produces two major virulence factors toxin A and B that inactivate Rho proteins by glucosylation of a pivotal threonine residue. Purified toxins induce reorganization of the cytoskeleton and cell death in colonic cells. Whether all toxin effects on target cells depend on catalytic glucosyltransferase activity is unclear at present. Thus, we conducted a proteome approach to compare the protein profile of target cells treated either with wild type toxin A (rTcdA wt) or with a catalytically inactive mutant toxin A (mutant rTcdA). Relative protein quantification was feasible using isotope-coded protein labeling techniques (ICPL) and mass spectrometry (LC-MALDI).ResultsAltogether we found a significant differential expression of thirty proteins after treatment with rTcdA wt or mutant rTcdA. Mutant rTcdA caused up-regulation of seven proteins and sixteen proteins were responsive to rTcdA wt after 5 h. Long-term effect of rTcdA wt on protein expression was the down-regulation of eleven proteins. Up- or down-regulation of several proteins was verified by western blot analysis confirming the MS results.ConclusionOur results indicate incubation time-dependent effects of the clostridial glucosylating toxin A on colonic cells. The rTcdA wt impact more cellular functions than actin cytoskeleton reorganization and apoptosis. Furthermore, these data give insight into glucosyltransferase independent effects of clostridial glucosylating toxins on target cells after short incubation time. Additionally, our data reveal pro-inflammatory and proliferative effects of mutant rTcdA after short-term incubation.

【 授权许可】

Unknown   
© Jochim et al; licensee BioMed Central Ltd. 2011. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.