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Interaction of diphtheria toxin and its active subunit, fragment A, with toxin-sensitive and toxin-resistant cells
Abstract:
Diphtheria toxin and purified fragment A, the active subunit of the toxin, were tested on toxin-sensitive and permeability class toxin-resistant cultured mammalian cells. Protein synthesis was inhibited to the same degree in sensitive or resistant cells by active concentrations of purified fragment A. In contrast, resistant cells required a concentration of whole toxin 5 to 6 logs greater than that required by sensitive cells to achieve the same degree of inhibition. On a molar basis, the toxicity of fragment A was equivalent to that of whole toxin on resistant cells. These results are evidence for the existence of two independent mechanisms for the entry of toxin or its active moiety into cells. One mechanism is a highly efficient, toxin-specific entry mechanism, involving surface receptor and fragment B-mediated association, and is active in sensitive cells. The other is a less efficient, nonspecific mechanism, probably related to endocytosis, which is operative in sensitive and resistant cells but is inapparent in sensitive cells when they are exposed to whole toxin because of the action of the specific mechanism.
Insights
Diphtheria toxin fragment A equally inhibits protein synthesis in sensitive and resistant cells, suggesting two distinct cellular entry mechanisms for the toxin. This reveals how cells take up diphtheria toxin.
Area of Science:
- Cell Biology
- Toxicology
- Molecular Biology
Background:
- Diphtheria toxin (DT) is a potent bacterial toxin that inhibits protein synthesis.
- Understanding DT cellular entry mechanisms is crucial for developing antitoxins and therapeutics.
Purpose of the Study:
- To investigate the mechanisms of DT entry into mammalian cells.
- To differentiate between toxin-specific and non-specific cellular uptake pathways.
Main Methods:
- Cultured toxin-sensitive and toxin-resistant mammalian cells were used.
- Protein synthesis inhibition was measured after exposure to purified DT fragment A and whole DT.
- Molar toxicity was compared between sensitive and resistant cell lines.
Main Results:
- Purified fragment A inhibited protein synthesis equally in both sensitive and resistant cells.
- Resistant cells required significantly higher concentrations of whole DT for equivalent inhibition compared to sensitive cells.
- Fragment A's molar toxicity was equivalent to whole DT in resistant cells.
Conclusions:
- Two distinct mechanisms mediate DT entry into mammalian cells.
- A specific, receptor-mediated pathway (involving fragment B) exists in sensitive cells.
- A less efficient, non-specific pathway (likely endocytosis) operates in both cell types but is masked by the specific pathway in sensitive cells.