Inhibition of protein synthesis in a cell-free system and Vero cells by okadaic acid, a diarrhetic shellfish toxin.
Okadaic acid, a diarrhetic shellfish toxin, is a potent promoter of tumors in mouse skin and a specific inhibitor of protein phosphatases 1 and 2A. In the present study, we investigated its effects on protein synthesis in Vero cells and rabbit reticulocyte lysate. Protein synthesis was inhibited by okadaic acid in Vero cells in a concentration-dependent manner (IC50 = 3.3 x 10(8) M-1). DNA synthesis was also inhibited by okadaic acid in Vero cells in a concentration-dependent manner (IC50 = 5.3 x 10(8) M-1). DNA synthesis inhibition in Vero cells occurred only after 8 h or longer. RNA synthesis was inhibited with an IC50 of 8.2 x 10(8) M-1. The time lag before DNA and RNA synthesis inhibition occurred was longer than the time lag before protein synthesis inhibition occurred in the same cells (4 h), indicating that protein synthesis is probably the main target and the first of okadaic acid's cytotoxic effects. Moreover, the inhibition of DNA and RNA synthesis is probably a consequence of the inhibition of protein synthesis. Since okadaic acid does not impair the uptake of the precursor of protein synthesis, it is assumed that the inhibition is due to a direct effect on one of the components of the protein synthesis machinery. We then used a cell-free system of rabbit reticulocyte lysate in which specific mRNA is translated into globin to ensure that protein synthesis is a direct target of okadaic acid in vitro. In rabbit reticulocyte lysate, okadaic acid inhibited protein synthesis in a concentration-dependent manner (IC50 = 6.3 x 10(12) M-1) with a correlation coefficient for percent inhibition values of r = .918. The molecular target of okadaic acid inside the cell whereby protein synthesis is inhibited remains to be discovered.
Matias
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WG
Laboratory of Toxicology and Applied Hygiene, Faculty of Pharmaceutical Sciences, University of Bordeaux, France.
Bonini
M
M
Creppy
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EE
D000818:Animals; D002452:Cell Count / Q000187:drug effects; D002474:Cell-Free System; D002522:Chlorocebus aethiops; D004261:DNA Replication / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000633:toxicity*; D004988:Ethers, Cyclic / Q000633:toxicity*; D066298:In Vitro Techniques; D007700:Kinetics; D008387:Marine Toxins / Q000633:toxicity*; D008974:Mollusca; D019319:Okadaic Acid; D014176:Protein Biosynthesis; D011506:Proteins / Q000187:drug effects*; D012313:RNA / Q000096:biosynthesis / Q000187:drug effects; D011817:Rabbits; D012156:Reticulocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D013997:Time Factors; D014709:Vero Cells
D004791:Enzyme Inhibitors; D004988:Ethers, Cyclic; D008387:Marine Toxins; D011506:Proteins; D019319:Okadaic Acid; D012313:RNA