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. 1995 Jun;63(6):2334–2340. doi: 10.1128/iai.63.6.2334-2340.1995

Isolation and characterization of a Clostridium botulinum C2 toxin-resistant cell line: evidence for possible involvement of the cellular C2II receptor in growth regulation.

G Fritz 1, P Schroeder 1, K Aktories 1
PMCID: PMC173306  PMID: 7768618

Abstract

Clostridium botulinum C2 toxin, which consists of the binding component C2II and the enzyme component C2I, acts on eukaryotic cells by selective ADP-ribosylation of G-actin. To obtain C2 toxin-resistant cells, we mutagenized CHO-K1 cells with N-nitroso-N-methylurea and selected for C2 resistance. Cells which survived the selection procedure with 50 ng of C2I and 100 ng of C2II per ml were obtained with a frequency of 30 x 10(-6). The colony-forming ability of CHO wild-type cells was reduced to 50% with 10 ng of C2I and 20 ng of C2II per ml. In contrast, the colony-forming ability of the isolated CHO mutant cells was not influenced by up to 200 ng of C2I and 400 ng of C2II per ml. Toxin-induced ADP-ribosylation of G-actin was not impaired in lysates of mutant cells. The C2 toxin-resistant phenotype remained sensitive to the cell-rounding activities of cytotoxins from C. perfringens (iota-toxin), C. novyi, C. difficile, and C. botulinum (C3) and to cytochalasin D. Binding of component C2II was impaired in resistant CHO cells, suggesting mutation of the toxin cell surface receptor. Serum factors protected wild-type cells against the cytotoxic effect of C2 toxin. Furthermore, the C2-resistant phenotype correlated with an increased serum dependency. The data suggest that the action of C. botulinum C2 toxin is mediated by its binding and uptake via a cell surface receptor which might be involved in growth regulation.

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Selected References

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