Multigenerational silencing dynamics control cell aging

Abstract

Cellular aging plays an important role in many diseases, such as cancers, metabolic syndromes, and neurodegenerative disorders. There has been steady progress in identifying aging-related factors such as reactive oxygen species and genomic instability, yet an emerging challenge is to reconcile the contributions of these factors with the fact that genetically identical cells can age at significantly different rates. Such complexity requires single-cell analyses designed to unravel the interplay of aging dynamics and cell-to-cell variability. Here we use microfluidic technologies to track the replicative aging of single yeast cells and reveal that the temporal patterns of heterochromatin silencing loss regulate cellular life span. We found that cells show sporadic waves of silencing loss in the heterochromatic ribosomal DNA during the early phases of aging, followed by sustained loss of silencing preceding cell death. Isogenic cells have different lengths of the early intermittent silencing phase that largely determine their final life spans. Combining computational modeling and experimental approaches, we found that the intermittent silencing dynamics is important for longevity and is dependent on the conserved Sir2 deacetylase, whereas either sustained silencing or sustained loss of silencing shortens life span. These findings reveal that the temporal patterns of a key molecular process can directly influence cellular aging, and thus could provide guidance for the design of temporally controlled strategies to extend life span.replicative aging | single-cell analysis | microfluidics | chromatin silencing | computational modeling C ellular aging is generally driven by the accumulation of genetic and cellular damage (1, 2). Although much progress has been made in identifying molecular factors that influence life span, what remains sorely missing is an understanding of how these factors interact and change dynamically during the aging process. This is in part because aging is a complex process wherein isogenic cells have various intrinsic causes of aging and widely different rates of aging. As a result, static population-based approaches could be insufficient to fully reveal sophisticated dynamic changes during aging. Recent developments in single-cell analyses to unravel the interplay of cellular dynamics and variability hold the promise to answer that challenge (3-5). Here we chose the replicative aging of yeast S. cerevisiae as a model and exploited quantitative biology technologies to study the dynamics of molecular processes that control aging at the single-cell level.

Quantitative analysis of the yeast pheromone pathway

“…However, there is a growing appreciation of the prevalence and importance of cell‐to‐cell variability (or “noise”) in biological processes. Such differences can arise from stochasticity in biochemical reactions, differences in the expression or activity of internal signalling components, age‐dependent accumulation of aggregated or damaged proteins, perturbations in membrane trafficking, and asynchronous progression through the cell cycle (Ansel et al, ; Becskei, Kaufmann, & van Oudenaarden, ; Colman‐Lerner et al, ; Elowitz, Levine, Siggia, & Swain, ; Fraser, Hirsh, Giaever, Kumm, & Eisen, ; Li et al, ; McAdams & Arkin, ; Paliwal et al, ; Pesce et al, ; Raser & O'Shea, ; Volfson et al, ; Yu et al, ). In that regard, fluorescent protein‐based reporters have been particularly useful because they permit quantitative measurements of induction in single, living cells over time.…”

Trajectories of Aging: How Systems Biology in Yeast Can Illuminate Mechanisms of Personalized Aging

“…Calorie restriction, accomplished by reducing the glucose present in the culture media, not only reduced the fraction of cells that followed the circular daughter trajectory, but also only increased the lifespan of cells with elliptical daughters (State 1). The authors suggest that the elliptical daughter state is likely to be related to increased rDNA circles or reduced rDNA silencing during aging, both of which have been previously implicated in yeast replicative aging through the Sir2‐dependent longevity pathway …”

“…Although the morphological changes reported by Jin et al. have been observed in other microfluidic systems, it is still important to determine whether these states are also seen in standard RLS assays, such as microdissection, and to determine whether specific microfluidic designs affect the penetrance of specific states.…”

An epigenetic landscape governs early fate decision in cellular aging

“…Using single-cell imaging technologies, we found that isogenic WT cells exhibit two different types of phenotypic changes during aging (6,7). About half of aging cells continuously produced daughters with a characteristic elongated morphology during later stages of lifespan.…”

“…A conserved lysine deacetylase Sir2, encoded by the best-studied longevity gene to date, mediates rDNA silencing (11). We recently used a GFP reporter inserted at the nontranscribed spacer region of the rDNA (rDNA-GFP) to track rDNA silencing in single cells (6). Since the expression of the reporter is repressed by silencing, increased fluorescence indicates a loss of silencing.…”