Figure 1.

Delocalization of telomeres VIII-left and XIV-left is dependent on expression of S-phase cyclins. (A) To determine the time of S-phase onset, DNA content was measured by FACS analysis in synchronized cultures of wild-type (GA-1985) and clb5 clb6 strains (HE4). Samples were taken every 10 min after release from α-factor synchronization. (B) Percentage of budded cells was analyzed in the synchronized cultures used in (A). (C, left panel) Microscopic image showing Tel VIII-left as a bright globular dot within the fluorescent nuclear rim. (Right panel) To quantify subnuclear positioning of a telomere, the area of the nucleus is measured and equally divided into three zones (Zone I, Zone II, and Zone III). Therefore, each zone occupies one-third of the area within the nuclear rim. In each cell, the telomere dot can be assigned to one of the three zones by measuring the distance from center of the dot to the periphery (indicated by the yellow line); a telomere is considered peripheral if it is located in Zone I. In a population of cells, a randomly positioned telomere will be present in Zones I, II, and III with equal frequency (∼33%), while an anchored telomere will be significantly enriched in Zone I. (D,E) Bar charts show the percentage of telomeres VIII-left and XIV-left in Zone I, II, or III. The asynchronous-growing cells were divided into four categories according to their bud size: unbudded, small-budded (<2 μm), medium-budded (2–2.75 μm), and large-budded (2.75–3.5 μm). The horizontal dashed line at 33% indicates the expected distribution of a randomly positioned telomere. Telomere positioning was observed and quantified in two independent strain isolates (error bars show SD). To determine statistical significance of telomere localization, P values from the χ² test were calculated by comparing the observed numbers in each zone with the expected random distribution in each population (significance values are indicated in Table 1).