Dataset Information

Two routes to senescence revealed by real-time analysis of telomerase-negative single lineages.

ABSTRACT: In eukaryotes, telomeres cap chromosome ends to maintain genomic stability. Failure to maintain telomeres leads to their progressive erosion and eventually triggers replicative senescence, a pathway that protects against unrestricted cell proliferation. However, the mechanisms underlying the variability and dynamics of this pathway are still elusive. Here we use a microfluidics-based live-cell imaging assay to investigate replicative senescence in individual Saccharomyces cerevisiae cell lineages following telomerase inactivation. We characterize two mechanistically distinct routes to senescence. Most lineages undergo an abrupt and irreversible switch from a replicative to an arrested state, consistent with telomeres reaching a critically short length. In contrast, other lineages experience frequent and stochastic reversible arrests, consistent with the repair of accidental telomere damage by Pol32, a subunit of polymerase δ required for break-induced replication and for post-senescence survival. Thus, at the single-cell level, replicative senescence comprises both deterministic cell fates and chaotic cell division dynamics.

SUBMITTER: Xu Z

PROVIDER: S-EPMC4503340 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Two routes to senescence revealed by real-time analysis of telomerase-negative single lineages.

Xu Zhou Z Fallet Emilie E Paoletti Camille C Fehrmann Steffen S Charvin Gilles G Teixeira Maria Teresa MT

Nature communications 20150709

In eukaryotes, telomeres cap chromosome ends to maintain genomic stability. Failure to maintain telomeres leads to their progressive erosion and eventually triggers replicative senescence, a pathway that protects against unrestricted cell proliferation. However, the mechanisms underlying the variability and dynamics of this pathway are still elusive. Here we use a microfluidics-based live-cell imaging assay to investigate replicative senescence in individual Saccharomyces cerevisiae cell lineage ...[more]

PMID: 26158780

Similar Datasets

Project description:BackgroundAccurate quantification of DNA using quantitative real-time PCR at low levels is increasingly important for clinical, environmental and forensic applications. At low concentration levels (here referring to under 100 target copies) DNA quantification is sensitive to losses during preparation, and suffers from appreciable valid non-detection rates for sampling reasons. This paper reports studies on a real-time quantitative PCR assay targeting a region of the human SRY gene over a concentration range of 0.5 to 1000 target copies. The effects of different sample preparation and calibration methods on quantitative accuracy were investigated.ResultsAt very low target concentrations of 0.5-10 genome equivalents (g.e.) eliminating any replicates within each DNA standard concentration with no measurable signal (non-detects) compromised calibration. Improved calibration could be achieved by eliminating all calibration replicates for any calibration standard concentration with non-detects ('elimination by sample'). Test samples also showed positive bias if non-detects were removed prior to averaging; less biased results were obtained by converting to concentration, including non-detects as zero concentration, and averaging all values. Tube plastic proved to have a strongly significant effect on DNA quantitation at low levels (p = 1.8 x 10(-4)). At low concentrations (under 10 g.e.), results for assays prepared in standard plastic were reduced by about 50% compared to the low-retention plastic. Preparation solution (carrier DNA or stabiliser) was not found to have a significant effect in this study.Detection probabilities were calculated using logistic regression. Logistic regression over large concentration ranges proved sensitive to non-detected replicate reactions due to amplification failure at high concentrations; the effect could be reduced by regression against log (concentration) or, better, by eliminating invalid responses.ConclusionUse of low-retention plastic tubes is advised for quantification of DNA solutions at levels below 100 g.e. For low-level calibration using linear least squares, it is better to eliminate the entire replicate group for any standard that shows non-detects reasonably attributable to sampling effects than to either eliminate non-detects or to assign arbitrary high Ct values. In calculating concentrations for low-level test samples with non-detects, concentrations should be calculated for each replicate, zero concentration assigned to non-detects, and all resulting concentration values averaged. Logistic regression is a useful method of estimating detection probability at low DNA concentrations.

Dataset Information

Two routes to senescence revealed by real-time analysis of telomerase-negative single lineages.

Publications

Two routes to senescence revealed by real-time analysis of telomerase-negative single lineages.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets