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In laboratory yeast strains with Sir2 and Fob1 function, wild-type NAD+ salvage is required for calorie restriction (CR) to extend replicative lifespan. CR does not significantly alter steady state levels of intracellular NAD+ metabolites. However, levels of Sir2 and Pnc1, two enzymes that sequentially convert NAD+ to nicotinic acid (NA), are up-regulated during CR. To test whether factors such as NA might be exported by glucose-restricted mother cells to survive later generations, we developed a replicative longevity paradigm in which mother cells are moved after 15 generations on defined media. The experiment reveals that CR mother cells lose the longevity benefit of CR when evacuated from their local environment to fresh CR media. Addition of NA or nicotinamide riboside (NR) allows a moved mother to maintain replicative longevity despite the move. Moreover, conditioned medium from CR-treated cells transmits the longevity benefit of CR to moved mother cells. Evidence suggests the existence of a longevity factor that is dialyzable but is neither NA nor NR, and indicates that Sir2 is not required for the longevity factor to be produced or to act. Data indicate that the benefit of glucose-restriction is transmitted from cell to cell in budding yeast, suggesting that glucose restriction may benefit neighboring cells and not only an individual cell.

Saccharomyces cerevisiae is calorie-restricted by lowering glucose from 2% to 0.5%. Under low glucose conditions, replicative lifespan is extended in a manner that depends on the NAD+-dependent protein lysine deacetylase Sir2 and NAD+ salvage enzymes. Because NAD+ is required for glucose utilization and Sir2 function, it was postulated that glucose levels alter the levels of NAD+ metabolites that tune Sir2 function. Though NAD+ precursor vitamins, which increase the levels of all NAD+ metabolites, can extend yeast replicative lifespan, glucose restriction does not significantly change the levels or ratios of intracellular NAD+ metabolites. To test whether glucose restriction affects protein copy numbers, we developed a technology that combines the measurement of Urh1 specific activity and quantification of relative expression between Urh1 and any other protein. The technology was applied to obtain the protein copy numbers of enzymes involved in NAD+ metabolism in rich and synthetic yeast media. Our data indicated that Sir2 and Pnc1, two enzymes that sequentially convert NAD+ to nicotinamide and then to nicotinic acid, are up-regulated by glucose restriction in rich media, and that Pnc1 alone is up-regulated in synthetic media while levels of all other enzymes are unchanged. These data suggest that production or export of nicotinic acid might be a connection between NAD+ and calorie restriction-mediated lifespan extension in yeast.

Saccharomyces cerevisiae is calorie-restricted by lowering glucose from 2% to 0.5%. Under low glucose conditions, replicative lifespan is extended in a manner that depends on the NAD+-dependent protein lysine deacetylase Sir2 and NAD+ salvage enzymes. Because NAD+ is required for glucose utilization and Sir2 function, it was postulated that glucose levels alter the levels of NAD+ metabolites that tune Sir2 function. Though NAD+ precursor vitamins, which increase the levels of all NAD+ metabolites, can extend yeast replicative lifespan, glucose restriction does not significantly change the levels or ratios of intracellular NAD+ metabolites. To test whether glucose restriction affects protein copy numbers, we developed a technology that combines the measurement of Urh1 specific activity and quantification of relative expression between Urh1 and any other protein. The technology was applied to obtain the protein copy numbers of enzymes involved in NAD+ metabolism in rich and synthetic yeast media. Our data indicated that Sir2 and Pnc1, two enzymes that sequentially convert NAD+ to nicotinamide and then to nicotinic acid, are up-regulated by glucose restriction in rich media, and that Pnc1 alone is up-regulated in synthetic media while levels of all other enzymes are unchanged. These data suggest that production or export of nicotinic acid might be a connection between NAD+ and calorie restriction-mediated lifespan extension in yeast. [ABSTRACT FROM AUTHOR]