The copper-sensing transcription factor Mac1, the histone deacetylase Hst1, and nicotinic acid regulate de novo NAD+ biosynthesis in budding yeast

Abstract

Edited by John M. Denu NADH (NAD ؉) is an essential metabolite involved in various cellular biochemical processes. The regulation of NAD ؉ metabolism is incompletely understood. Here, using budding yeast (Saccharomyces cerevisiae), we established an NAD ؉ intermediate-specific genetic system to identify factors that regulate the de novo branch of NAD ؉ biosynthesis. We found that a mutant strain (mac1⌬) lacking Mac1, a copper-sensing transcription factor that activates copper transport genes during copper deprivation, exhibits increases in quinolinic acid (QA) production and NAD ؉ levels. Similar phenotypes were also observed in the hst1⌬ strain, deficient in the NAD ؉-dependent histone deacetylase Hst1, which inhibits de novo NAD ؉ synthesis by repressing BNA gene expression when NAD ؉ is abundant. Interestingly, the mac1⌬ and hst1⌬ mutants shared a similar NAD ؉ metabolism-related gene expression profile, and deleting either MAC1 or HST1 de-repressed the BNA genes. ChIP experiments with the BNA2 promoter indicated that Mac1 works with Hst1-containing repressor complexes to silence BNA expression. The connection of Mac1 and BNA expression suggested that copper stress affects de novo NAD ؉ synthesis, and we show that copper stress induces both BNA expression and QA production. Moreover, nicotinic acid inhibited de novo NAD ؉ synthesis through Hst1-mediated BNA repression, hindered the reuptake of extracellular QA, and thereby reduced de novo NAD ؉ synthesis. In summary, we have identified and characterized novel NAD ؉ homeostasis factors. These findings will expand our understanding of the molecular basis and regulation of NAD ؉ metabolism. NAD ϩ and its reduced form NADH are primary redox carriers in cellular metabolism. NAD ϩ is also a cosubstrate in protein modifications, such as protein deacetylation mediated by the sirtuins (Sir2 family proteins) and ADP-ribosylation mediated by the poly(ADP-ribose) polymerases. These protein modifications contribute to the maintenance and regulation of chromatin structure, DNA repair, circadian rhythm, metabolic responses, and life span (1-4). NAD ϩ is also an NADP ϩ precursor, which, like NAD ϩ , is carefully balanced with its reduced form NADPH to maintain a favorable redox state. Aberrant NAD ϩ metabolism is associated with a number of diseases, including diabetes, cancer, and neuron degeneration (2, 3, 5-11). Administration of NAD ϩ precursors, such as nicotinamide mononucleotide (NMN), 2 nicotinamide (NAM), nicotinic acid riboside, and nicotinamide riboside (NR), has been shown to ameliorate deficiencies related to aberrant NAD ϩ metabolism in yeast, mouse, and human cells (3, 5-10, 12-15). However, the molecular mechanisms underlying the beneficial effects of NAD ϩ precursor supplementation are not yet completely understood. The NAD ϩ pool is maintained by multiple NAD ϩ biosynthesis pathways, which are conserved from bacteria to humans. Depending on the cell types, growth conditions, and availability of specific NAD ϩ precursors, one pathway may dominate the others. ...

Vitamin requirements and biosynthesis in Saccharomyces cerevisiae

“…Hst1 does not bind the DNA directly but interacts with Rfm1 and Sum1 to form a repressor complex. Mac1, which was previously characterized as a copper‐sensing transcription factor, has been shown to also be involved in regulation of BNA genes, together with Hst1 (Bedalov et al, ; James Theoga Raj et al, ; Laurenson & Rine, ; McCord et al, ). When NA is abundantly available, NA salvage metabolism is preferred over use of the de novo biosynthetic pathway, which is repressed by Hst1 (Bedalov et al, ; James Theoga Raj et al, ).…”

“…Mac1, which was previously characterized as a copper‐sensing transcription factor, has been shown to also be involved in regulation of BNA genes, together with Hst1 (Bedalov et al, ; James Theoga Raj et al, ; Laurenson & Rine, ; McCord et al, ). When NA is abundantly available, NA salvage metabolism is preferred over use of the de novo biosynthetic pathway, which is repressed by Hst1 (Bedalov et al, ; James Theoga Raj et al, ). In S. cerevisiae , NAD + metabolism is regulated together with phosphate and purine nucleotide metabolism, although the exact mechanisms remain uncharacterized (Lu & Lin, ; Pinson, Ceschin, Saint‐Marc, & Daignan‐Fornier, ).…”

NAD+ Metabolism and Regulation: Lessons From Yeast

“…Therefore, the cell must maintain these metabolites and their flux in a controlled manner. We do not fully understand all the mechanisms by which the cell can sense and tune these metabolites, but some known NAD + homeostasis regulatory mechanisms include transcriptional control, feedback inhibition, nutrient sensing, and enzyme or metabolite compartmentalization [1,31,34,35,[37][38][39][40][41][42].…”

“…Because Hst1 activity is dependent on NAD + , NAD + depletion results in transcription activation of the de novo pathway. A recent study showed that the copper-sensing transcription factor Mac1 appeared to work in concert with Hst1 to repress BNA genes [34].…”

“…In yeast, mutants carrying single and multiple deletions of NAD + pathway components and special defined growth conditions that pinpoint certain pathways are relatively easy to obtain. Several NAD + homeostasis factors were uncovered in recent studies using NAD + precursor-specific genetic screens [31,[34][35][36]. Given the interconnections among NAD + biosynthesis pathways and cellular processes, identification and studying additional NAD + homeostasis factors are required to elucidate the regulation of cellular NAD + metabolism.…”