Selective inhibition of cytokinesis in sea urchin embryos by low concentrations of stypoldione, a marine natural product that reacts with sulfhydryl groups.

Stypoldione is a marine natural product that inhibits cells division in marine embryos and in mammalian cell cultures. The mechanism responsible for the ability of the compound to inhibit cell division is not known. The compound was found in early studies to inhibit polymerization of tubulin into microtubules in vitro, which lead to the suggestion that inhibition of microtubule polymerization in cells might be responsible for the ability of the compound to inhibit cell division. More recently, stypoldione was found to react covalently with the sulfhydryl groups of a number of proteins including tubulin and with sulfhydryl groups of peptides and small molecules. Thus, stypoldione could potentially react with a large number of cellular targets. In the present study, we have examined the effects of stypoldione on the organization of microtubules and chromatin in cells, in relation to the ability of the compound to inhibit cell division. We used indirect immunofluorescence light microscopy of fixed and stained sea urchin embryos during the first and second divisions after fertilization, with stains specific for tubulin and DNA. We found that stypoldione exerted qualitatively different effects on cell division and microtubule organization and function at different concentrations. At the lowest effective concentrations, 5-10 microM, stypoldione selectively inhibited cytokinesis. Mitotic division occurred normally, usually with no discernible perturbation of microtubule organization or function, and cells became multinuclear. At somewhat higher concentrations, 20-40 microM, stypoldione blocked embryos before streak stage of the first division and, although microtubules were present, their organization was perturbed and they often formed unusual "spiral aster" arrays. At 80 microM and above, microtubules in blocked cells were largely absent. Thus, stypoldione uncouples cytokinesis from mitosis at the lowest effective concentrations and, although it can disrupt microtubules at relatively higher concentrations, it inhibits cell division at the lowest effective concentrations by a selective action on cytokinesis through a mechanism that does not appear to involve disassembly of microtubules.