Depletion of Htl1 causes decreased Nbp1 and Ndc1 at SPBs. (a) Nbp1-GFP was expressed from the ADH1 promoter in a kap123Δ strain or the HTL1 shut-off strain containing Spc42-mCherry, and summed projections of a representative large-budded cell are shown. Bar, 3 µm. (b) Wild-type and HTL1 shut-off strains expressing Nbp1-GFP and carrying a high-copy plasmid expressing NBP1 were grown in medium containing glucose for the indicated times. A summed projection of a representative large-budded cell for each time point in the HTL1 shut-off is shown. Bar, 3 µm. Corresponding flow cytometry histograms can be found in Fig. S6 a. (c) The mean GFP intensity per pixel in the whole cell was measured for 100 cells at the indicated times by using summed intensity projections from panel b. Error bars represent one SD from the mean. (d) Levels of Nbp1-GFP at >200 SPBs were measured in wild-type and HTL1 shut-off strains carrying a high-copy plasmid expressing NBP1 at the indicated times during growth in glucose. The ratio of Nbp1-GFP to Spc42-mCherry at the SPBs is plotted. The black bars represent the medians. (e) Levels of Ndc1-GFP at SPBs or NPCs were measured in the HTL1 shut-off strain at the indicated times. The ratio of SPB-associated Ndc1-GFP to Spc42-mCherry for at least 200 SPBs is shown on the left. The amount of Ndc1-GFP at the NPC cells is displayed on the right. The black bars represent the medians. Corresponding flow cytometry histograms can be found in Fig. S6 d. The P values were calculated using a two-tailed Mann-Whitney test.