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. 1999 Aug;10(8):2771–2785. doi: 10.1091/mbc.10.8.2771

Figure 7.

Figure 7

Premature recruitment of Plo1 to the SPBs in stf1.1 mutant cells. (A) An asynchronous stf1.1 culture was fixed and stained using HN184 antibodies and DAPI. Plo1 localized to the SPB of small interphase cells. The cells in A have shrunk after processing for immunofluorescence. This is because a very short fixation regime is used for Plo1 staining, and the cell structure has contracted in the 2 d before this image was captured. All statistical analysis throughout this article were conducted on freshly stained cells before shrinkage occurred. (B) Asynchronous wild-type IH365 and stf1.1 cells were processed for immunofluoresence using HN184 antibodies and DAPI. The presence or absence of Plo1 staining was noted, and the frequency of a positive score was plotted as a functionof the cells’ length. This was carried out for a minimum of 30 cells for each length category. Plo1 localizes to smaller cells and a higher proportion of stf1.1 cells than wild-type. (C) A culture of stf1.1 cells was synchronized by elutrient centrifugation. Samples were taken every 20 min and mixed with asynchronous cdc7. A20 mutant cells that had been at the restrictive temperature for 4–6 h. These samples were stained with HN184 antibodies and DAPI. The inclusion of the septation mutant provided an internal control for the quality of sample preparation in the sample because the long multinucleate cdc7. A20 cells were easily distinguishable from the shorter stf1.1 cells. Thus the ratio of Plo1 SPB association in stf1.1 relative to cdc7. A20 staining gives a measure of Plo1 association with the stf1.1 SPB in interphase. The level of SPB association in cdc7. A20 remained constant throughout the experiment, but the relative staining on the SPBs of stf1.1 cells varied. Two hundred cells of each strain were scored at each time point. The ratio of Plo1 association in stf1.1 cells relative to cdc7. A20 cells is presented (circles) alongside the septation index (open squares). Although this complex experiment was only performed once with a synchronous culture, data from the analysis of mixed asynchronous populations result in similar conclusions (see RESULTS).