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Temporal separation of protein toxin translocation from processing events.
1Laboratory of Molecular Biology, National Institute of Mental Health, Bethesda, Maryland 20892.
The Journal of Biological Chemistry
|December 5, 1987
Summary
The transport of ricin, modeccin, diphtheria toxin (DT), and Pseudomonas exotoxin A to the cytosol is the rate-limiting step in protein synthesis inhibition. Acidification is crucial for establishing translocation probability but not for the translocation itself.
Area of Science:
- Cell Biology
- Molecular Toxicology
- Biochemistry
Background:
- Cellular intoxication by toxins like ricin, modeccin, diphtheria toxin (DT), and Pseudomonas exotoxin A involves receptor binding, cytoplasmic transport, and inhibition of protein synthesis.
- Previous studies indicated two cell subpopulations during toxin intoxication: one active in protein synthesis and another inhibited.
Purpose of the Study:
- To correlate autoradiographic data with protein synthesis decline rates across multiple toxins.
- To elucidate the rate-limiting steps and cellular mechanisms involved in toxin-induced protein synthesis inhibition.
Main Methods:
- Intoxication of Vero cells with ricin, modeccin, DT, and Pseudomonas exotoxin A.
- Kinetic analysis of protein synthesis decline.
- Autoradiographic analysis to identify cell subpopulations with varying protein synthesis activity.
Main Results:
- All four toxins induced a lag phase followed by a pseudo first-order decrease in protein synthesis.
- Autoradiography revealed two distinct cell subpopulations during intoxication: actively synthesizing and inhibited cells.
- Cytosolic transport of toxins was identified as the rate-limiting step, with translocation releasing sufficient toxin to rapidly halt protein synthesis.
Conclusions:
- Cytosolic transport is the rate-limiting step for protein synthesis inhibition by these toxins.
- Acidification is required for establishing translocation probability during the lag phase, not for the translocation event itself.
- A kinetic model for DT intoxication, from endocytosis to translocation, was proposed, highlighting shared cellular bases with protein synthesis decline.