Genetic Manipulation of Glycogen Allocation Affects Replicative Lifespan in E. coli.
Bottom Line: We found that certain changes in the regulation of the carbohydrate metabolism can affect aging.These results demonstrate how manipulations of nutrient allocation can lead to the exclusion of the chromosome and limit replicative lifespan in E. coli, and illustrate how mutations can have phenotypic effects that are specific for cells with old poles.This raises the question how bacteria can avoid the accumulation of such mutations in their genomes over evolutionary times, and how they can achieve the long replicative lifespans that have recently been reported.
Affiliation: Biozentrum, University of Basel, Switzerland.
In bacteria, replicative aging manifests as a difference in growth or survival between the two cells emerging from division. One cell can be regarded as an aging mother with a decreased potential for future survival and division, the other as a rejuvenated daughter. Here, we aimed at investigating some of the processes involved in aging in the bacterium Escherichia coli, where the two types of cells can be distinguished by the age of their cell poles. We found that certain changes in the regulation of the carbohydrate metabolism can affect aging. A mutation in the carbon storage regulator gene, csrA, leads to a dramatically shorter replicative lifespan; csrA mutants stop dividing once their pole exceeds an age of about five divisions. These old-pole cells accumulate glycogen at their old cell poles; after their last division, they do not contain a chromosome, presumably because of spatial exclusion by the glycogen aggregates. The new-pole daughters produced by these aging mothers are born young; they only express the deleterious phenotype once their pole is old. These results demonstrate how manipulations of nutrient allocation can lead to the exclusion of the chromosome and limit replicative lifespan in E. coli, and illustrate how mutations can have phenotypic effects that are specific for cells with old poles. This raises the question how bacteria can avoid the accumulation of such mutations in their genomes over evolutionary times, and how they can achieve the long replicative lifespans that have recently been reported.
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Mentions: We thus used a second approach where we analyzed the intracellular localization of a fusion protein of the glycogen synthase GlgA to the green fluorescent protein (GFP). It has been reported that GlgA cocrystallizes with oligosaccharides , indicating that determining the localization of GlgA can inform us about the spatial distribution of glycogen in the cell. The GlgA-GFP fusion protein was preferentially localizing at the old poles of csrA mutant cells (Fig 3A, S3 Movie), and also, yet to a lesser extent, of wild type cells (S4 Movie). More importantly, the GlgA-GFP signal accumulates at old poles of csrA mutant cells over consecutive divisions (Fig 3A), and after the last division event, GlgA-GFP appeared to be distributed across the whole cytoplasm (Fig 3A). We quantified the distribution of GlgA-GFP for mother and daughter cells of first, second last, and last divisions in csrA mutant cells (Fig 3B). The results support a gradual accumulation of glycogen at the old pole that eventually leads to cells that are filled with glycogen (S5 Fig). Analysis of the GlgA-GFP concentration inside a cell and the cell’s length after division showed that both traits are predictive of whether the observed division is this cell’s final division or not (S6 Fig). Additionally, a cell’s GlgA-GFP concentration and cell size when it emerges as a new-pole cell is predictive of its total replicative potential; the brighter and smaller cells are ‘at birth’ when they emerge as a new-pole cell, the lower the total replicative lifespan they will reach (S7 Fig). As a control, we observed cells expressing GFP alone, without fusion to another protein, under control of the promoter for the ribosomal protein RpsM; we found no evidence for the distribution of GPF being different between young and old pole daughter cells, showing that the effect observed with GlgA- GFP is not due to passive accumulation of GFP molecules at the cell poles (S8 Fig).