Figure 2. Nucleolar contraction upon nutrient depletion requires condensin and Hmo1.

(A) Cell density of wild-type cells growing in rich medium (YPD) over time. Cells were seeded at low density (8.5 × 10⁵ cells/ml), OD₆₀₀ = 0.2. Arrow indicates exit from log phase. (B–C) Relative nucleolar volume from the same time course. Samples were taken at the indicated time points, fixed, and stained for Nop1 and nuclear GFP (tetR-NLS-GFP) to determine nucleolar and nuclear volume, respectively. (B) Representative images of Nop1 (red) and nuclear GFP (green) staining. (C) Relative nucleolar volume was determined for 25 cells for each time point and was normalized to the 4-hr time point. (D) Repeat-specific URA3 expression before and after treatment with rapamycin or ethanol (mock) was measured by qPCR. Values are normalized against UBC6 and repeat 49 per treatment. Error bars indicate s.d. of 3 independent experiments, each analyzed in triplicate. (E) Representative IF images of cells stained for Nop1 (red) and DAPI (blue) to label the nucleolus and non-nucleolar chromatin. Scale bar = 1μm. (F) Relative nucleolar volume compared to nuclear GFP normalized to wild type log phase at 23°C; n=25 for each condition. (G) Representative IF images of hmo1Δ mutants grown at 30°C in synthetic complete medium. Cells were stained for Nop1 (red) and DAPI (blue). Arrow indicates DAPI-negative gap in hmo1Δ mutants. (H) Relative nucleolar volume as in (F); n=25. Nucleolar volume was normalized to wild type log phase. (I) Distance between the centroids of the nucleolus and the non-nucleolar chromatin in wild type and hmo1Δ mutants as depicted in the schematic (top); n=35. Significance tests: (D) Welch two-sample t-test; (C, F, H, I) Wilcoxon signed-rank test. See also Figure S2.