Low concentrations of the toxin ophiobolin A lead to an arrest of the cell cycle and alter the intracellular partitioning of glutathione between the nuclei and cytosol

Ophiobolin A, a tetracyclic sesterpenoid produced by phytopathogenic fungi, is responsible for catastrophic losses incrop yield but its mechanism of action is not understood. The effects of ophiobolin A were therefore investigated on thegrowth and redox metabolism of Tobacco Bright Yellow-2 (TBY-2) cell cultures by applying concentrations of the toxinthat did not promote cell death. At concentrations between 2–5 μM, ophiobolin A inhibited growth and proliferation ofthe TBY-2 cells, which remained viable. Microscopic and cytofluorimetric analyses showed that ophiobolin A treatmentcaused a rapid decrease in mitotic index, with a lower percentage of the cells at G1 and increased numbers of cells atthe S/G2 phases. Cell size was not changed following treatment suggesting that the arrest of cell cycle progression wasnot the result of a block on cell growth. The characteristic glutathione redox state and the localization of glutathione inthe nucleus during cell proliferation were not changed by ophiobolin A. However, subsequent decreases in glutathioneand the re-distribution of glutathione between the cytoplasm and nuclei after mitosis occurring in control cells, as wellas the profile of glutathionylated proteins, were changed in the presence of the toxin. The profile of poly ADP-ribosylatedproteins were also modified by ophiobolin A. Taken together, these data provide evidence of the mechanism of ophiobolinA action as a cell cycle inhibitor and further demonstrate the link between nuclear glutathione and the cell cycle regulation,suggesting that glutathione-dependent redox controls in the nuclei prior to cell division are of pivotal importance.