Figure 6.

Phosphorylation sites within NLS are required for efficient release from a late mitotic block. (A) Mutation of NLS phosphorylation sites impaired mitotic exit after reversal of cell cycle arrest. CDC14 and cdc14-(PS1,2A) cultures were placed at 37°C for 2 h to achieve a late mitotic cdc15-2 arrest and returned to permissive temperature (25°C). Clb2p and Cdc28 levels were followed by immunoblot analysis. (B) Cell aliquots from A were stained with DAPI and evaluated by fluorescence microscopy. Combined DIC and DAPI fluorescence images are shown. (C, left) Clb2 protein detected in A was quantified and normalized to Cdc28 levels. Relative units of fluorescence intensity were plotted for each time point. (right) The frequency of large-budded cells (n > 200) with segregated nuclei (B) was plotted for each time point. WT, wild type. Bars, 2 µm.