Figure 5

Wiskostatin does not affect cellular ATP levels. A. Cellular ATP levels in wiskostatin treated cells. HeLa cells were synchronised in prometaphase with nocodazole and released after shake-off in presence of vehicle (control), 10 mM NaN₃ + 6 mM deoxyglucose (ΔATP) or 10 μM wiskostatin (wiskostatin). After indicated time points cells were harvested and lysed in boiling water for 10 minutes. Cellular ATP contents were determined on clarified lysates using ATP determination kit (see Methods). Cellular ATP level at time point 0 from each treatment was defined as 100%. Graph represents mean of three independent experiments. Error bars represent SEM. ATP depletion delays mitosis. B. HeLa cells synchronised and treated as previously described were harvested at indicated time points and DNA content analysed by flow cytometry. C. In similarly treated cells cyclin B1 and γ-tubulin levels were determined by immunoblot analysis using specific antibodies.