Evolution of Distinct Responses to Low NAD+ Stress by Rewiring the Sir2 Deacetylase Network in Yeasts

Abstract

Evolutionary adaptation increases the fitness of a species in its environment. It can occur through rewiring of gene regulatory networks, such that an organism responds appropriately to environmental changes. We investigated whether sirtuin deacetylases, which repress transcription and require NAD+ for activity, serve as transcriptional rewiring points that facilitate the evolution of potentially adaptive traits. If so, bringing genes under the control of sirtuins could enable organisms to mount appropriate responses to stresses that decrease NAD+ levels. To explore how the genomic targets of sirtuins shift over evolutionary time, we compared two yeast species, Saccharomyces cerevisiae and Kluyveromyces lactis, that display differences in cellular metabolism and life cycle timing in response to nutrient availability. We identified sirtuin-regulated genes through a combination of chromatin immunoprecipitation and RNA expression. In both species, regulated genes were associated with NAD+ homeostasis, mating, and sporulation, but the specific genes differed. In addition, regulated genes in K. lactis were associated with other processes, including utilization of nonglucose carbon sources, detoxification of arsenic, and production of the siderophore pulcherrimin. Consistent with the species-restricted regulation of these genes, sirtuin deletion affected relevant phenotypes in K. lactis but not S. cerevisiae. Finally, sirtuin-regulated gene sets were depleted for broadly conserved genes, consistent with sirtuins regulating processes restricted to a few species. Taken together, these results are consistent with the notion that sirtuins serve as rewiring points that allow species to evolve distinct responses to low NAD+ stress.

A single chromosome strain of S. cerevisiae exhibits diminished ethanol metabolism and tolerance

“…Further, the majority of genes that are upregulated in sirtuin deletion strains are also signi cantly upregulated in our reference strain during growth on ethanol (Additional File 1: Fig. S8) 27,31 . These repressed genes include genes encoding enzymes involved in non-fermentable carbon source growth including CIT2, PDC6, YAT1, and transcription factors including ADR1, which in uences post-diauxic gene expression 27,31 .…”

“…This rescue result indicates that the genes necessary for ethanol growth and metabolism are present in SY14, but that one or more of these is repressed. Previous works suggest that sirtuins connect the metabolic state of yeast to gene expression responses by sensing NAD+ 27,28 , and deletion of SIR2 has been shown to enhance ethanol catabolism 29 and gluconeogenesis 30 . Further, the majority of genes that are upregulated in sirtuin deletion strains are also signi cantly upregulated in our reference strain during growth on ethanol (Additional File 1: Fig.…”

“…S8) 27,31 . These repressed genes include genes encoding enzymes involved in non-fermentable carbon source growth including CIT2, PDC6, YAT1, and transcription factors including ADR1, which in uences post-diauxic gene expression 27,31 . These data suggest that the switch-like change in gene expression upon glucose depletion in S. cerevisiae may be driven by alleviation of sirtuin-mediated silencing of speci c genes.…”

A single chromosome strain of S. cerevisiae exhibits diminished ethanol metabolism and tolerance

“…This rescue result indicates that the genes necessary for ethanol growth and metabolism are present in SY14, but that one or more of these is repressed. Previous works suggest that sirtuins connect the metabolic state of yeast to gene expression responses by sensing NAD+ [ 28 , 29 ], and deletion of SIR2 has been shown to enhance ethanol catabolism [ 30 ] and gluconeogenesis [ 31 ]. Further, the majority of genes that are upregulated in sirtuin deletion strains are also significantly upregulated in our reference strain during growth on ethanol (Additional File 1 : Fig.…”

“…Further, the majority of genes that are upregulated in sirtuin deletion strains are also significantly upregulated in our reference strain during growth on ethanol (Additional File 1 : Fig. S8) [ 28 , 32 ]. These repressed genes include genes encoding enzymes involved in non-fermentable carbon source growth including CIT2 , PDC6, YAT1, and transcription factors including ADR1 , which influences post-diauxic gene expression [ 28 , 32 ].…”