Fig. 1
From: Spatial reorganization of telomeres in long-lived quiescent cells
Massive telomere reorganization upon carbon source exhaustion. a Growth curve for S. cerevisiae grown in rich glucose-based liquid medium. Yeast cells grown in medium containing glucose divide exponentially, mainly perform glycolysis, and release ethanol into the medium. When glucose becomes limiting (roughly after 12 hours in the conditions used in this study; see “Materials and methods”) the cells undergo a major metabolic transition called “diauxic shift”, during which they stop fermentation and start aerobic utilization of ethanol (respiration phase). After this transition, cells divide slowly and become more resistant to different stresses. Once ethanol is exhausted and no other carbon source is available, around 7 days, the culture enters the stationary phase (SP). At this stage, the majority of the cells are in a quiescent state. b Representative fluorescent images of the telomere-associated protein Rap1 tagged with green fluorescent protein (GFP). Overnight wild-type (WT) “yAT1684” liquid cultures were diluted to 0.2 OD600nm/ml and images were acquired after 5 hours (1 OD600nm/ml, fermentation phase), 2 days (28 OD600nm/ml, respiration phase) and 7 days (40 OD600nm/ml, stationary phase). c Quantification of the distribution of intensity and number of foci of Rap1-GFP images from experiment shown in (b) with our in-house software. Pie charts represent percentages of cells with 0 (white), 1 (red), 2 (orange), 3 (green) and 4 (blue) foci. Box plots: white = fermentation (Ferm), light gray = respiration (Resp), dark grey = stationary (Stat). Median (line) and mean (cross) are indicated. For each condition, more than 1000 cells were analyzed. Statistical tests were carried out using the Mann–Whitney non-parametric test (****p < 0.0001; ***p < 0.001; **0.001 < p < 0.01; *0.01 < p < 0.05; ns = p > 0). d Colocalization of telomeres with Rap1 foci. ImmunoFISH with Y’ probes was performed on a WT strain yAT1684 at SP. e Representative fluorescent images of the telomere-associated protein Rap1 tagged with GFP in SP WT and sir3∆ cells. f Rap1-GFP hypercluster localization relative to the nuclear pore. Two-color z-stack images were acquired on a WT strain yAT2407 expressing Rap1-yemRFP and the GFP tagged nucleoporin 49 (Nup49-GFP) during SP. The localization of the Rap1-yemRFP hypercluster in one of the three equal concentric zones of the nucleus was scored in the focal plane. This experiment was repeated twice and for each experiment >100 nuclei with a hyper-cluster were analyzed