Fig. 2. The effects of analogue on cdc2-as division and kinase activity. (A,B) The indicated cells were grown in YE medium to early exponential phase and analogue was added to the indicated final concentrations; the controls received an equal volume of DMSO. Samples were removed at the indicated times and analysed by FACS. (C) A protein extract was prepared from cdc2-as 10 minutes after release from the analogue arrest. As a control, protein extracts were prepared from cdc2⁺ cdc25-22 arrest-release-synchronised cells (20 minute time point, when cells are in early mitosis and Cdc2p activity is high) and assayed. Analogue, or an equivalent volume of solvent, was added to the extracts at the indicated concentrations, and an H1 kinase assay was performed. The upper panel shows a scanned autoradiograph of the phosphorylated H1. The middle panels show the Coomassie-Blue-stained histone H1 and the bottom panels show a western blot of the extracts used for the assay, probed with TAT-1 antibody as loading control. Note that the kinase activity is reduced by addition of analogue to the cdc2-as extract, but not the cdc2⁺ extract.