Yeast chronological aging is regarded as a model for aging of mammalian post-mitotic cells. It refers to changes occurring in stationary phase cells over a relatively long period of time. How long these cells can survive in such a non-dividing state defines the chronological lifespan. Several factors influence cell survival including two well known normal by-products of yeast glucose fermentation such as ethanol and acetic acid. In fact, the presence in the growth medium of these C2 compounds has been shown to limit the chronological lifespan. In the chronological aging paradigm, a pro-aging role has also emerged for the deacetylase Sir2, the founding member of the Sirtuin family, whose loss of function increases the depletion of extracellular ethanol by an unknown mechanism. Here, we show that lack of Sir2 strongly influences carbon metabolism. In particular, we point out a more efficient acetate utilization which in turn may have a stimulatory effect on ethanol catabolism. This correlates with an enhanced glyoxylate/gluconeogenic flux which is fuelled by the acetyl-CoA produced from the acetate activation. Thus, when growth relies on a respiratory metabolism such as that on ethanol or acetate, SIR2 inactivation favors growth. Moreover, in the chronological aging paradigm, the increase in the acetate metabolism implies that sir2Δ cells avoid acetic acid accumulation in the medium and deplete ethanol faster; consequently pro-aging extracellular signals are reduced. In addition, an enhanced gluconeogenesis allows replenishment of intracellular glucose stores which may be useful for better long-term cell survival.

In Saccharomyces cerevisiae, the chronological lifespan (CLS) is defined as the length of time that a population of nondividing cells can survive in stationary phase. In this phase, cells remain metabolically active, albeit at reduced levels, and responsive to environmental signals, thus simulating the postmitotic quiescent state of mammalian cells. Many studies on the main nutrient signaling pathways have uncovered the strong influence of growth conditions, including the composition of culture media, on CLS. In this context, two byproducts of yeast glucose fermentation, ethanol and acetic acid, have been proposed as extrinsic proaging factors. Here, we report that ethanol and acetic acid, at physiological levels released in the exhausted medium, both contribute to chronological aging. Moreover, this combined proaging effect is not due to a toxic environment created by their presence but is mainly mediated by the metabolic pathways required for their utilization as carbon/energy sources. In addition, measurements of key enzymatic activities of the glyoxylate cycle and gluconeogenesis, together with respiration assays performed in extreme calorie restriction, point to a long-term quiescent program favoured by glyoxylate/gluconeogenesis flux contrary to a proaging one based on the oxidative metabolism of ethanol/acetate via TCA and mitochondrial respiration.

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ACH1 encodes a mitochondrial enzyme of Saccharomyces cerevisiae endowed with CoA-transferase activity. It catalyzes the CoASH transfer from succinyl-CoA to acetate generating acetyl-CoA. It is known that ACH1 inactivation results in growth defects on media containing acetate as a sole carbon and energy source which are particularly severe at low pH. Here, we show that chronological aging ach1Δ cells which accumulate a high amount of extracellular acetic acid display a reduced chronological lifespan. The faster drop of cell survival is completely abrogated by alleviating the acid stress either by a calorie restricted regimen that prevents acetic acid production or by transferring chronologically aging mutant cells to water. Moreover, the short-lived phenotype of ach1Δ cells is accompanied by reactive oxygen species accumulation, severe mitochondrial damage, and an early insurgence of apoptosis. A similar pattern of endogenous severe oxidative stress is observed when ach1Δ cells are cultured using acetic acid as a carbon source under acidic conditions. On the whole, our data provide further evidence of the role of acetic acid as cell-extrinsic mediator of cell death during chronological aging and highlight a primary role of Ach1 enzymatic activity in acetic acid detoxification which is important for mitochondrial functionality.

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