Protein acetylation microarray reveals that NuA4 controls key metabolic target regulating gluconeogenesis

Area of Science:

• Biochemistry
• Molecular Biology
• Cell Biology

Background:

• Histone acetyltransferases (HATs) and histone deacetylases (HDACs) regulate critical cellular functions via nonhistone substrates in metazoans.
• In Saccharomyces cerevisiae, only chromatin-associated nonhistone substrates were previously known.
• The essential nucleosome acetyltransferase of H4 (NuA4) complex plays a key role in cellular regulation.

Purpose of the Study:

• To identify and characterize nonchromatin substrates of the NuA4 complex in yeast.
• To investigate the role of Pck1p acetylation in glucose metabolism and cellular lifespan.
• To explore the evolutionary conservation of Pck1p regulation by HATs/HDACs.

Main Methods:

• Yeast proteome microarrays for substrate identification.
• Tandem mass spectrometry for determining acetylation sites.
• In vitro and in vivo enzymatic assays.
• Analysis of yeast chronological lifespan and glucose production in human cells.

Main Results:

• Identified numerous nonchromatin substrates for the NuA4 complex.
• Determined acetylation sites (Lys19 and 514) on phosphoenolpyruvate carboxykinase (Pck1p).
• Acetylation at Lys514 is essential for Pck1p enzymatic activity and yeast growth on nonfermentable carbon sources.
• Sir2p deacetylated Pck1p both in vitro and in vivo.
• Pck1p activity loss impaired lifespan extension under water starvation.
• Human Pck1 acetylation and glucose production in HepG2 cells depend on TIP60 (ESA1 homolog).

Conclusions:

• The NuA4 complex regulates glucose metabolism and lifespan through acetylation of the metabolic enzyme Pck1p.
• Pck1p acetylation is a conserved regulatory mechanism impacting cellular physiology and lifespan.
• This study reveals a novel link between histone acetyltransferases, metabolic enzymes, and aging.