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

ABSTRACT: 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.

SUBMITTER: James Theoga Raj C

PROVIDER: S-EPMC6462523 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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

James Theoga Raj Christol C Croft Trevor T Venkatakrishnan Padmaja P Groth Benjamin B Dhugga Gagandeep G Cater Timothy T Lin Su-Ju SJ

The Journal of biological chemistry 20190213 14

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 (mac1Δ) lacking Mac1, a copper-sensing transcription factor tha ...[more]

PMID: 30760525

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Project description:Intracellular pathogens residing within macrophage phagosomes are challenged with recognizing the phagosomal environment and appropriately responding to changing host defense strategies imposed in this organelle. One such phagocyte defense is the restriction of available copper as a form of nutritional immunity during the adaptive immune response to fungal pathogens. The intracellular fungal pathogen Histoplasma capsulatum adapts to this decreased copper through upregulation of the Ctr3 copper transporter. In this study, we show that Histoplasma recognizes the characteristic low-copper phagosomal environment of activated macrophages through the copper-dependent transcriptional regulator Mac1. Multiple cis-acting regulatory sequences in the CTR3 promoter control upregulation of CTR3 transcription under low-copper conditions, and the loss of Mac1 function prevents induction of Ctr3 under low-copper conditions. During adaptive immunity, this loss of copper sensing by Mac1 attenuates Histoplasma virulence more severely than loss of Ctr3 alone, indicating that Mac1 controls the expression of additional mechanisms important for pathogenesis. Transcriptional profiling of Histoplasma yeasts identified a small regulon of Mac1-dependent genes, with the most strongly regulated genes encoding proteins linked to copper, iron, and zinc homeostasis and defenses against reactive oxygen (iron-requiring catalase [CatB] and superoxide dismutase [Sod4]). Accordingly, the loss of Mac1 function increased sensitivity to copper and iron restriction and blocked low-copper-induced expression of extracellular catalase activity. Thus, Mac1-mediated sensing of low-copper signals to Histoplasma yeasts their residence within the activated macrophage phagosome, and in response, Histoplasma yeasts produce factors, including non-copper-dependent factors, to combat the enhanced defenses of activated macrophages. IMPORTANCE Histoplasma capsulatum is a fungal pathogen that survives and grows within host macrophages. For successful infection, Histoplasma must sense and adapt to a dynamic intracellular environment over the course of an infection. We demonstrate that the copper-dependent transcription factor, Mac1, enables Histoplasma sensing of low copper that characterizes the phagosome environment of activated macrophages. Histoplasma recognition of this state leads not only to upregulation of copper acquisition mechanisms but also to other non-copper-related pathogenesis strategies, including scavenging of other metals and detoxification of reactive oxygen produced by host cells. The limited set of genes regulated by Histoplasma Mac1 compared to those of other fungal pathogens suggests a response that has been tailored specifically for Histoplasma's life inside the phagosome. Thus, low levels of phagosomal copper serve as a signal to Histoplasma, enabling responses to the enhanced antimicrobial defenses resulting from immune activation of macrophages.

Dataset Information

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

Publications

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

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