Microtubule-dependent cell cycle regulation is implicated in the G2 phase of Hydra cells

Interstitial cells of Hydra attenuata, from which nerve cells and nematocytes (stinging cells) differentiate, were arrested in either metaphase or G2 by different concentrations of the microtubule-depolymerizing agent nocodazole. At a concentration of 1.4 nM-nocodazole, a large number of cells were arrested in metaphase. However, at concentrations of 2 nM-nocodazole and above most of the cells were arrested at a distinct point in G2 several hours before mitosis. After removal of the 2 nM-nocodazole block, 75% of the cells entered the next cell cycle about 10 h later. To our knowledge this is the first time that cells have been synchronized by arresting them in the G2 phase. Visualization of Hydra microtubules with a tubulin monoclonal antibody and immunofluorescent staining showed that the very low concentrations of nocodazole used for cell cycle arrest were indeed affecting microtubule structures. Spindles and stem cell microtubules disappeared at 0.8-1 nM-nocodazole, followed by nerve microtubules (about 2 nM), cnidocil microtubules (10 nM) and finally by nematocyte microtubules (34 nM). Taken together, these data strongly indicate a microtubule-dependent mechanism of cell cycle regulation in the G2 phase.