A functional link between NAD + homeostasis and N-terminal protein acetylation in Saccharomyces cerevisiae

J Biol Chem. 2018 Feb 23;293(8):2927-2938. doi: 10.1074/jbc.M117.807214. Epub 2018 Jan 9.

Abstract

Nicotinamide adenine dinucleotide (NAD+) is an essential metabolite participating in cellular redox chemistry and signaling, and the complex regulation of NAD+ metabolism is not yet fully understood. To investigate this, we established a NAD+-intermediate specific reporter system to identify factors required for salvage of metabolically linked nicotinamide (NAM) and nicotinic acid (NA). Mutants lacking components of the NatB complex, NAT3 and MDM20, appeared as hits in this screen. NatB is an Nα-terminal acetyltransferase responsible for acetylation of the N terminus of specific Met-retained peptides. In NatB mutants, increased NA/NAM levels were concomitant with decreased NAD+ We identified the vacuolar pool of nicotinamide riboside (NR) as the source of this increased NA/NAM. This NR pool is increased by nitrogen starvation, suggesting NAD+ and related metabolites may be trafficked to the vacuole for recycling. Supporting this, increased NA/NAM release in NatB mutants was abolished by deleting the autophagy protein ATG14 We next examined Tpm1 (tropomyosin), whose function is regulated by NatB-mediated acetylation, and Tpm1 overexpression (TPM1-oe) was shown to restore some NatB mutant defects. Interestingly, although TPM1-oe largely suppressed NA/NAM release in NatB mutants, it did not restore NAD+ levels. We showed that decreased nicotinamide mononucleotide adenylyltransferase (Nma1/Nma2) levels probably caused the NAD+ defects, and NMA1-oe was sufficient to restore NAD+ NatB-mediated N-terminal acetylation of Nma1 and Nma2 appears essential for maintaining NAD+ levels. In summary, our results support a connection between NatB-mediated protein acetylation and NAD+ homeostasis. Our findings may contribute to understanding the molecular basis and regulation of NAD+ metabolism.

Keywords: NAD biosynthesis; cell metabolism; metabolic regulation; metabolism; yeast genetics.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Acetylation
  • Acetyltransferases / chemistry
  • Acetyltransferases / genetics
  • Acetyltransferases / metabolism
  • Autophagy-Related Proteins / genetics
  • Autophagy-Related Proteins / metabolism
  • Gene Deletion
  • Genes, Reporter
  • Homeostasis
  • Immunoprecipitation
  • Isoenzymes / chemistry
  • Isoenzymes / genetics
  • Isoenzymes / metabolism
  • Models, Molecular*
  • Mutation
  • N-Terminal Acetyltransferase B / chemistry
  • N-Terminal Acetyltransferase B / genetics
  • N-Terminal Acetyltransferase B / metabolism*
  • NAD / metabolism*
  • Nicotinamide-Nucleotide Adenylyltransferase / chemistry
  • Nicotinamide-Nucleotide Adenylyltransferase / genetics
  • Nicotinamide-Nucleotide Adenylyltransferase / metabolism*
  • Protein Interaction Domains and Motifs
  • Protein Multimerization
  • Protein Processing, Post-Translational*
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Saccharomyces cerevisiae / enzymology
  • Saccharomyces cerevisiae / growth & development
  • Saccharomyces cerevisiae / metabolism*
  • Saccharomyces cerevisiae Proteins / chemistry
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Tropomyosin / genetics
  • Tropomyosin / metabolism

Substances

  • ATG 14 protein, S cerevisiae
  • Autophagy-Related Proteins
  • Isoenzymes
  • MDM20 protein, S cerevisiae
  • Nat3 protein, S cerevisiae
  • Recombinant Proteins
  • Saccharomyces cerevisiae Proteins
  • TPM1 protein, S cerevisiae
  • Tropomyosin
  • NAD
  • Acetyltransferases
  • NatB protein, S cerevisiae
  • N-Terminal Acetyltransferase B
  • Nicotinamide-Nucleotide Adenylyltransferase
  • nicotinic acid mononucleotide adenylyltransferase